U.S. patent application number 10/456861 was filed with the patent office on 2004-07-29 for 3 (5) -heteroaryl substituted pyrazoles as p38 kinase inhibitors.
Invention is credited to Clare, Michael, Geng, Lifeng, Hanson, Gunnar J., Naraian, Ashok S., Partis, Richard A., Stealey, Michael A., Weier, Richard M..
Application Number | 20040147579 10/456861 |
Document ID | / |
Family ID | 22181538 |
Filed Date | 2004-07-29 |
United States Patent
Application |
20040147579 |
Kind Code |
A1 |
Naraian, Ashok S. ; et
al. |
July 29, 2004 |
3 (5) -heteroaryl substituted pyrazoles as p38 kinase
inhibitors
Abstract
A class of pyrazole derivatives is described for use in treating
p38 kinase mediated disorders. Compounds of particular interest are
defined by Formula I 1 wherein R.sup.1, R.sup.2, Ar.sup.1 and
HetAr.sup.2 are as described in the specification.
Inventors: |
Naraian, Ashok S.; (Ballwin,
MO) ; Clare, Michael; (US) ; Geng, Lifeng;
(San Marcos, CA) ; Hanson, Gunnar J.; (Skokie,
IL) ; Partis, Richard A.; (Grayslake, IL) ;
Stealey, Michael A.; (Libertyville, IL) ; Weier,
Richard M.; (Lake Bluff, IL) |
Correspondence
Address: |
HARNESS, DICKEY, & PIERCE, P.L.C
7700 BONHOMME, STE 400
ST. LOUIS
MO
63105
US
|
Family ID: |
22181538 |
Appl. No.: |
10/456861 |
Filed: |
June 6, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10456861 |
Jun 6, 2003 |
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10024071 |
Dec 18, 2001 |
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6579873 |
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Current U.S.
Class: |
514/406 ;
548/364.1 |
Current CPC
Class: |
C07D 401/04 20130101;
C07D 401/14 20130101 |
Class at
Publication: |
514/406 ;
548/364.1 |
International
Class: |
A61K 031/415; C07D
403/02 |
Claims
What is claimed is:
1. A compound of Formula I 28wherein R.sup.1 is selected from
hydrido, alkyl, cycloalkyl, alkenyl, alkynyl, heterocyclyl,
cycloalkylalkylene, haloalkyl, hydroxyalkyl, aralkyl, alkoxyalkyl,
mercaptoalkyl, alkylthicalkylene, amino, alkylamino, arylamino,
aminoalkyl, alkylaminoalkylene, heterocyclylalkylene,
aminocarbonylalkylene, and alkylaminocarbonylalkylene; and R.sup.2
is selected from hydrido, alkyl, alkenyl, alkynyl, heterocyclyl,
haloalkyl, heterocyclylalkyl, amino, alkylamino, aminoalkyl,
alkoxy, alkylthio, carboxy, alkoxycarbonyl, carboxyalkyl,
aminocarbonylamino, alkylaminocarbonylamino, alkylsulfonyl,
aminosulfonyl, alkylsulfonylamino, aminosulfonylamino,
alkylaminosulfonylamino, and alkynylamino; wherein the heterocyclyl
and heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, alkoxy, aryloxy, aralkoxy,
heterocyclyl, haloalkyl, amino, cyano, and hydroxy; and Ar.sup.1 is
aryl optionally substituted with one or more radicals independently
selected from halo, alkyl, alkenyl, alkynyl, alkoxy, alkenoxy,
alkyldioxy, alkylthio, alkylsulfinyl, alkylsulfonyl, amino,
aminocarbonyl, cyano, alkoxycarbonyl, formyl, aminosulfonyl,
alkylamino, nitro, arylamino, alkylcarbonylamino, halosulfonyl,
aminoalkyl, and haloalkyl; and HetAr.sup.2 is pyridinyl,
pyrimidinyl or quinolinyl optionally substituted with one or more
radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, heterocyclyl, alkoxy, aralkoxy,
haloalkyl, amino, cyano, aralkyl, alkylamino, cycloalkylamino,
cycloalkenylamino, arylamino, alkynylamino, and aralkylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
2. A compound of claim 1 wherein R.sup.1 is selected from hydrido,
lower alkyl, lower cycloalkyl, lower cycloalkylalkylene, lower
haloalkyl, lower hydroxyalkyl, lower alkenyl, lower alkynyl, lower
heterocyclyl, lower aralkyl, lower alkoxyalkyl, lower
mercaptoalkyl, lower alkylthioalkylene, amino, lower alkylamino,
lower arylamino, lower aminoalkyl, lower alkylaminoalkylene, lower
heterocyclylalkylene, lower aminocarbonylalkylene, and lower
alkylaminocarbonylalkylene; and R.sup.2 is selected from hydrido,
lower alkyl, lower alkenyl, lower alkynyl, lower haloalkyl, lower
heterocyclyl, lower heterocyclylalkylene, amino, lower alkylamino,
lower alkynylamino, lower aminoalkyl, lower alkylthio, lower
carboxy, lower alkoxycarbonyl, lower carboxyalkyl, lower
aminocarbonylamino, lower alkylaminocarbonylamino, lower
alkylsulfonyl, lower aminosulfonyl, lower alkylsulfonylamino, lower
aminosulfonylamino, and lower alkylaminosulfonylamino, wherein the
heterocyclyl and heterocyclylalkyl groups are optionally
substituted with one or more radicals independently selected from
lower alkylthio, lower alkylsulfonyl, lower alkylsulfinyl, halo,
lower alkyl, lower alkoxy, aryloxy, lower heterocyclyl, lower
haloalkyl, amino, and cyano; and Ar.sup.1 is selected from phenyl,
biphenyl, and naphthyl, wherein Ar.sup.1 is optionally substituted
with one or more radicals independently selected from lower
alkylthio, lower alkylsulfonyl, aminosulfonyl, halo, lower alkyl,
lower alkenyl, lower alkynyl, lower alkylsulfinyl, cyano, lower
alkoxycarbonyl, aminocarbonyl, formyl, lower alkylcarbonylamino,
lower haloalkyl, lower alkoxy, lower alkenyloxy, lower alkyldioxy,
amino, lower alkylamino, lower aminoalkyl, arylamino, nitro, and
halosulfonyl; and HetAr.sup.2 is pyridinyl or pyrimidinyl
optionally substituted with one or more radicals independently
selected from lower alkylthio, lower alkylsulfonyl, lower
alkylsulfinyl, halo, lower alkyl, lower heterocyclyl, lower alkoxy,
lower aralkoxy, lower haloalkyl, amino, cyano, lower aralkyl, lower
alkylamino, lower cycloalkylamino, lower arylamino, lower
alkynylamino, and lower aralkylamino; or a
pharmaceutically-acceptable salt or tautomer thereof.
3. A Compound of claim 2 wherein R.sup.1 is selected from hydrido,
methyl, ethyl, isopropyl, tert-butyl, isobutyl, trichloroethyl,
pentafluoroethyl, heptafluoropropyl, difluoroethyl, difluoropropyl,
dichloroethyl, dichloropropyl, vinyl, allyl, ethynyl, propargyl,
morpholinyl, piperidinyl, piperazinyl, benzyl, phenylethyl,
morpholinomethyl, morpholinoethyl, pyrrolidinylmethyl,
piperazinylmethyl, piperidinylmethyl, pyridinylmethyl,
thienylmethyl, methoxymethyl, ethoxymethyl, amino, methylamino,
dimethylamino, phenylamino, methylaminomethyl, dimethylaminomethyl,
methylaminoethyl, dimethylaminoethyl, cyclopropyl, cyclopentyl,
cyclohexyl, cyclohexylmethyl, hydroxymethyl, hydroxyethyl,
methylthio, and methylthiomethyl; and R.sup.2 is selected from
hydrido, methyl, ethyl, propyl, isopropyl, tert-butyl, isobutyl,
fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl,
dichloromethyl, trichloromethyl, pentafluoroethyl,
heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl,
difluoroethyl, difluoropropyl, dichloroethyl, dichloropropyl,
amino, N-methylamino, N,N-dimethylamino, ethynylamino,
propargylamino, piperidinyl, piperazinyl, morpholinomethyl,
pyrrolidinylmethyl, piperazinylmethyl, piperidinylmethyl,
pyridinylmethyl, thienylmethyl, thiazolylmethyl, oxazolylmethyl,
pyrimidinylmethyl, quinolylmethyl, isoquinolinylmethyl,
imidazolylmethyl, benzimidazolylmethyl, furylmethyl,
pyrazinylmethyl, aminocarbonylamino, methylaminocarbonylamino,
dimethylaminocarbonylamino, ethylaminocarbonylamino,
diethylaminocarbonylamino, methylsulfonylamino, ethylsulfonylamino,
aminosulfonylamino, methylaminosulfonylamino,
dimethylaminosulfonylamino, ethylaminosulfonylamino, and
diethylaminosulfonylamino; and Ar.sup.1 is selected from phenyl,
biphenyl, and naphthyl, wherein Ar.sup.1 is optionally substituted
with one or more radicals independently selected from methylthio,
methylsulfinyl, methylsulfonyl, fluoro, chloro, bromo,
aminosulfonyl, methyl, ethyl, isopropyl, tert-butyl, isobutyl,
cyano, methoxycarbonyl, ethoxycarbonyl, aminocarbonyl,
methylcarbonylamino, trifluoromethyl, difluoromethyl, fluoromethyl,
trichloromethyl, dichloromethyl, chloromethyl, allyl, vinyl,
ethynyl, propargyl, methoxy, ethoxy, propyloxy, n-butoxy, amino,
methylamino, ethylamino, dimethylamino, diethylamino, aminomethyl,
aminoethyl, N-methyl, N-phenylamino, phenylamino, diphenylamino,
nitro, and chlorosulfonyl; and HetAr.sup.2 is selected from
pyridinyl and pyrimidinyl, wherein HetAr.sup.2 is optionally
substituted with one or more radicals independently selected from
methylthio, methylsulfinyl, methylsulfonyl, fluoro, chloro, bromo,
methyl, ethyl, isopropyl, tert-butyl, isobutyl, methoxyl, ethoxyl,
phenoxyl, benzoxyl, phenethyl, trifluoromethyl, fluoromethyl,
difluoromethyl, amino, benzylamino, propargylamino,
cyclopropylamino, cyclobutylamino, cyclopentylamino, and cyano; or
a pharmaceutically-acceptable salt or tautomer thereof.
4. A compound of claim 3 wherein R.sup.1 is hydrido, methyl, ethyl,
hydroxyethyl, propargyl, dimethylaminoethyl or morpholinoethyl; and
R.sup.2 is selected from hydrido, methyl, ethyl, amino,
aminocarbonylamino, methylaminocarbonylamino, methylsulfonylamino,
aminosulfonylamino, and methylaminosulfonylamino; and Ar.sup.1 is
phenyl optionally substituted with one or more radicals
independently selected from methylthio, methylsulfinyl,
methylsulfonyl, fluoro, chloro, bromo, aminosulfonyl, methyl,
ethyl, isopropyl, tert-butyl, isobutyl, cyano, methoxycarbonyl,
ethoxycarbonyl, aminocarbonyl, methylcarbonylamino,
trifluoromethyl, difluoromethyl, fluoromethyl, trichloromethyl,
dichloromethyl, chloromethyl, methoxy, ethoxy, propyloxy, n-butoxy,
amino, methylamino, ethylamino, dimethylamino, diethylamino,
aminomethyl, aminoethyl, N-methyl, N-phenylamino, phenylamino,
diphenylamino, nitro, and chlorosulfonyl; and HetAr.sup.2 is
optionally substituted with one or more radicals independently
selected from methylthio, methylsulfinyl, methylsulfonyl, fluoro,
chloro, bromo, methyl, ethyl, isopropyl, tert-butyl, isobutyl,
methoxyl, ethoxyl, phenoxyl, benzoxyl, trifluoromethyl,
fluoromethyl, difluoromethyl, amino, propargylamino, and cyano; or
a pharmaceutically-acceptable salt or a tautomer thereof.
5. A compound of claim 4 wherein R.sup.1 is hydrido or methyl; and
R.sup.2 is hydrido or methyl; and Ar.sup.1 is phenyl which is
optionally substituted with one or more radicals independently
selected fluoro, chloro, methyl, ethyl, trifluoromethyl, methoxy,
ethoxy, dimethylamino, and nitro; and HetAr.sup.2 is optionally
substituted with one or more radicals independently selected from
methyl, chloro, fluoro, and trifluoromethyl; or a
pharmaceutically-acceptable salt or tautomer thereof.
6. A compound of claim 1 wherein R.sup.2 is hydrido.
7. A compound of claim 2 wherein R.sup.2 is hydrido.
8. A compound of claim 3 wherein R.sup.2 is hydrido.
9. A compound of claim 4 wherein R.sup.2 is hydrido.
10. A compound of claim 5 wherein R.sup.2 is hydrido.
11. A compound of claim 1 wherein HetAr.sup.2 is optionally
substituted pyridinyl.
12. A compound of claim 2 wherein HetAr.sup.2 is optionally
substituted pyridinyl.
13. A compound of claim 3 wherein HetAr.sup.2 is optionally
substituted pyridinyl.
14. A compound of claim 4 wherein HetAr.sup.2 is optionally
substituted pyridinyl.
15. A compound of claim 5 wherein HetAr.sup.2 is optionally
substituted pyridinyl.
16. A compound of claim 1 wherein R.sup.2 is hydrido, Ar.sup.1 is
optionally substituted phenyl, and HetAr.sup.2 is optionally
substituted pyridinyl.
17. A compound of claim 2 wherein R.sup.2 is hydrido, Ar.sup.1 is
optionally substituted phenyl, and HetAr.sup.2 is optionally
substituted pyridinyl.
18. A compound of claim 3 wherein R.sup.2 is hydrido, Ar.sup.1 is
optionally substituted phenyl, and HetAr.sup.2 is optionally
substituted pyridinyl.
19. A compound of claim 4 wherein R.sup.2 is hydrido and
HetAr.sup.2 is optionally substituted pyridinyl.
20. A compound of claim 5 wherein R.sup.2 is hydrido and
HetAr.sup.2 is optionally substituted pyridinyl.
21. A compound of claim 4 selected from the compounds, their
tautomers and their pharmaceutically acceptable salts, of the group
consisting of 4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridine;
4-(4-fluorophenyl)-5-(4-py- ridinyl)-1H-pyrazol-3-amine;
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyraz-
ol-3-yl]methanesulfonamide;
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyraz-
ol-3-yl]-N'-methylsulfamide;
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazo- l-3-yl]urea;
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]sulfamide- ;
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine;
N-[4-(4-fluorophenyl)-5-14-pyridinyl)-1H-pyrazol-3-yl]-N'-methylurea;
4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine;
4-[4-(4-fluorophenyl)-1-m- ethyl-1H-pyrazol-3-yl]pyridine;
4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyra- zole-1-ethanol;
4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazol-
e-1-ethanamine;
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]e-
thyl]morpholine; 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine;
1-methyl-4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]]piperi-
dine; and
1-methyl-4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-1-y-
l]]piperidine.
22. A compound of Formula I 29wherein R.sup.1 is selected from
hydrido, lower alkyl, lower cycloalkyl, lower cycloalkylalkylene,
lower haloalkyl, lower hydroxyalkyl, lower alkynyl, lower aralkyl,
lower alkoxyalkyl, lower mercaptoalkyl, lower alkylthioalkylene,
amino, lower alkylamino, lower arylamino, lower aminoalkyl, lower
alkylaminoalkylene, lower heterocyclylalkylene, lower
aminocarbonylalkylene, and lower alkylaminocarbonylalkylene; and
R.sup.2 is selected from hydrido, lower alkyl, lower cycloalkyl,
lower cycloalkenyl, aryl selected from phenyl and biphenyl, lower
aralkyl, lower haloalkyl, lower heterocyclyl, lower
heterocyclylalkylene, amino, lower arylamino, lower alkylamino,
lower alkynylamino, lower aminocarbonylamino, lower
alkylaminocarbonylamino, sulfonyl, lower alkylsulfonylamino, lower
aminosulfonylamino, and lower alkylaminosulfonylamino, wherein the
cycloalkyl, aryl and heterocyclyl groups are optionally substituted
with one or more radicals independently selected from lower
alkylthio, lower alkylsulfonyl, lower alkylsulfinyl, halo, lower
alkyl, lower alkoxy, aryloxy, lower heterocyclyl, lower haloalkyl,
amino, and cyano; and Ar.sup.1 is phenyl optionally substituted
with one or more radicals independently selected from halo, lower
alkyl, lower alkenyl, lower alkynyl, lower alkoxy, lower alkylthio,
lower alkylsulfinyl, lower alkylsulfonyl, amino, aminocarbonyl,
cyano, lower alkoxycarbonyl, formyl, aminosulfonyl, lower
alkylamino, nitro, lower arylamino, lower alkylcarbonylamino,
halosulfonyl, lower aminoalkyl, and lower haloalkyl; and
HetAr.sup.2 is pyridinyl optionally substituted with one or more
radicals independently selected from lower alkylthio, lower
alkylsulfonyl, lower alkylsulfinyl, halo, lower alkyl, lower
heterocyclyl, lower alkoxy, lower aralkoxy, lower haloalkyl, amino,
cyano, lower aralkyl, lower alkylamino, lower cycloalkylamino,
lower aralkylamino, and lower arylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
23. A compound of claim 22 wherein R.sup.1 is hydrido, methyl,
hydroxyethyl, dimethylaminoethyl, propargyl, or morpholinoethyl;
and R.sup.2 is selected from hydrido, methyl, ethyl, amino,
aminocarbonylamino, methylaminocarbonylamino, methylsulfonylamino,
aminosulfonylamino, and methylaminosulfonylamino;
24. A compound of claim 22 wherein R.sup.2 is hydrido.
25. A compound of claim 22 wherein Ar.sup.1 is phenyl substituted
with one or more halogen radicals.
26. A compound of claim 22 wherein R.sup.2 is hydrido and Ar.sup.1
is phenyl substituted with one or more halogen radicals.
27. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 1; or a pharmaceutically salt
or tautomer thereof.
28. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 2; or a pharmaceutically salt
or tautomer thereof.
29. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 3; or a pharmaceutically salt
or tautomer thereof.
30. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 4; or a pharmaceutically salt
or tautomer thereof.
31. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 5; or a pharmaceutically salt
or tautomer thereof.
32. A pharmaceutical composition comprising a
therapeutically-effective amount of a compound, said compound
selected from the compounds of claim 21; or a pharmaceutically salt
or tautomer thereof.
33. A method of treating a TNF mediated disorder, said method
comprising treating the subject having or susceptible to such
disorder with a therapeutically-effective amount of a compound of
Formula I 30wherein R.sup.1 is selected from hydrido, alkyl,
cycloalkyl, alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene,
haloalkyl, hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthioalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, heterocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkylene; and R.sup.2 is selected from
hydrido, alkyl, alkenyl, alkynyl, heterocyclyl, haloalkyl,
heterocyclylalkyl, amino, alkylamino, aminoalkyl, alkoxy,
alkylthio, carboxy, alkoxycarbonyl, carboxyalkyl,
aminocarbonylamino, alkylaminocarbonylamino, alkylsulfonyl,
aminosulfonyl, alkylsulfonylamino, aminosulfonylamino,
alkylaminosulfonylamino, and alkynylamino; wherein the heterocyclyl
and heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, alkoxy, aryloxy, aralkoxy,
heterocyclyl, haloalkyl, amino, cyano, and hydroxy; and Ar.sup.1 is
aryl optionally substituted with one or more radicals independently
selected from halo, alkyl, alkenyl, alkynyl, alkoxy, alkenoxy,
alkyldioxy, alkylthio, alkylsulfinyl, alkylsulfonyl, amino,
aminocarbonyl, cyano, alkoxycarbonyl, formyl, aminosulfonyl,
alkylamino, nitro, arylamino, alkylcarbonylamino, halosulfonyl,
aminoalkyl, and haloalkyl; and HetAr.sup.2 is pyridinyl,
pyrimidinyl or quinolinyl optionally substituted with one or more
radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, heterocyclyl, alkoxy, aralkoxy,
haloalkyl, amino, cyano, aralkyl, alkylamino, cycloalkylamino,
cycloalkenylamino, arylamino, alkynylamino, and aralkylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
34. A method of treating a p38 kinase mediated disorder, said
method comprising treating the subject having or susceptible to
such disorder with a therapeutically-effective amount of a compound
of Formula I 31wherein R.sup.1 is selected from hydrido, alkyl,
cycloalkyl, alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene,
haloalkyl, hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthicalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, heterocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkyle- ne; and R.sup.2 is selected from
hydrido, alkyl, alkenyl, alkynyl, heterocyclyl, haloalkyl,
heterocyclylalkyl, amino, alkylamino, aminoalkyl, alkoxy,
alkylthio, carboxy, alkoxycarbonyl, carboxyalkyl,
aminocarbonylamino, alkylaminocarbonylamino, alkylsulfonyl,
aminosulfonyl, alkylsulfonylamino, aminosulfonylamino,
alkylaminosulfonylamino, and alkynylamino; wherein the heterocyclyl
and heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, alkoxy, aryloxy, aralkoxy,
heterocyclyl, haloalkyl, amino, cyano, and hydroxy; and Ar.sup.1 is
aryl optionally substituted with one or more radicals independently
selected from halo, alkyl, alkenyl, alkynyl, alkoxy, alkenoxy,
alkyldioxy, alkylthio, alkylsulfinyl, alkylsulfonyl, amino,
aminocarbonyl, cyano, alkoxycarbonyl, formyl, aminosulfonyl,
alkylamino, nitro, arylamino, alkylcarbonylamino, halosulfonyl,
aminoalkyl, and haloalkyl; and HetAr.sup.2 is pyridinyl,
pyrimidinyl or quinolinyl optionally substituted with one or more
radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, heterocyclyl, alkoxy, aralkoxy,
haloalkyl, amino, cyano, aralkyl, alkylamino, cycloalkylamino,
cycloalkenylamino, arylamino, alkynylamino, and aralkylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
35. A method of treating inflammation, said method comprising
treating the subject having or susceptible to such condition with a
therapeutically-effective amount of a compound of Formula I
32wherein R.sup.1 is selected from hydrido, alkyl, cycloalkyl,
alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene, haloalkyl,
hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthioalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, heterocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkylene; and R.sup.2 is selected from
hydrido, alkyl, alkenyl, alkynyl, heterocyclyl, haloalkyl,
heterocyclylalkyl, amino, alkylamino, aminoalkyl, alkoxy,
alkylthio, carboxy, alkoxycarbonyl, carboxyalkyl,
aminocarbonylamino, alkylaminocarbonylamino, alkylsulfonyl,
aminosulfonyl, alkylsulfonylamino, aminosulfonylamino,
alkylaminosulfonylamino, and alkynylamino; wherein the heterocyclyl
and heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, alkoxy, aryloxy, aralkoxy,
heterocyclyl, haloalkyl, amino, cyano, and hydroxy; and Ar.sup.1 is
aryl optionally substituted with one or more radicals independently
selected from halo, alkyl, alkenyl, alkynyl, alkoxy, alkenoxy,
alkyldioxy, alkylthio, alkylsulfinyl., alkylsulfonyl, amino,
aminocarbonyl, cyano, alkoxycarbonyl, formyl, aminosulfonyl,
alkylamino, nitro, arylamino, alkylcarbonylamino, halosulfonyl,
aminoalkyl, and haloalkyl; and HetAr.sup.2 is pyridinyl,
pyrimidinyl or quinolinyl optionally substituted with one or more
radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, heterocyclyl, alkoxy, aralkoxy,
haloalkyl, amino, cyano, aralkyl, alkylamino, cycloalkylamino,
cycloalkenylamino, arylamino, alkynylamino, and aralkylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
36. A method of treating arthritis, said method comprising treating
the subject having or susceptible to such condition with a
therapeutically-effective amount of a compound of Formula I
33wherein R.sup.1 is selected from hydrido, alkyl, cycloalkyl,
alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene, haloalkyl,
hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthioalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, heterocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkylene; and R.sup.2 is selected from
hydrido, alkyl, alkenyl, alkynyl, heterocyclyl, haloalkyl,
heterocyclylalkyl, amino, alkylamino, aminoalkyl, alkoxy,
alkylthio, carboxy, alkoxycarbonyl, carboxyalkyl,
aminocarbonylamino, alkylaminocarbonylamino, alkylsulfonyl,
aminosulfonyl, alkylsulfonylamino, aminosulfonylamino,
alkylaminosulfonylamino, and alkynylamino; wherein the heterocyclyl
and heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, alkoxy, aryloxy, aralkoxy,
heterocyclyl, haloalkyl, amino, cyano, and hydroxy; and Ar.sup.1 is
aryl optionally substituted with one or more radicals independently
selected from halo, alkyl, alkenyl, alkynyl, alkoxy, alkenoxy,
alkyldioxy, alkylthio, alkylsulfinyl, alkylsulfonyl, amino,
aminocarbonyl, cyano, alkoxycarbonyl, formyl, aminosulfonyl,
alkylamino, nitro, arylamino, alkylcarbonylamino, halosulfonyl,
aminoalkyl, and haloalkyl; and HetAr.sup.2 is pyridinyl,
pyrimidinyl or quinolinyl optionally substituted with one or more
radicals independently selected from alkylthio, alkylsulfonyl,
alkylsulfinyl, halo, alkyl, heterocyclyl, alkoxy, aralkoxy,
haloalkyl, amino, cyano, aralkyl, alkylamino, cycloalkylamino,
cycloalkenylamino, arylamino, alkynylamino, and aralkylamino; or a
pharmaceutically-acceptable salt or a tautomer thereof.
37. The method of claim 33 wherein the TNF mediated disorder is
selected from the group of disorders consisting of bone resorption,
graft vs. host reaction, atherosclerosis, arthritis,
osteoarthritis, rheumatoid arthritis, gout, psoriasis, topical
inflammatory disease state, adult respiratory distress syndrome,
asthma, chronic pulmonary inflammatory disease, cardiac reperfusion
injury, renal reperfusion injury, thrombus, glomerulonephritis,
Crohn's disease, ulcerative colitis, inflammatory bowel disease and
cachexia.
38. The method of claim 33 wherein the TNF mediated disorder is
inflammation.
39. The method of claim 33 wherein the TNF mediated disorder is
arthritis.
40. The method of claim 33 wherein the TNF mediated disorder is
asthma.
41. The method of claim 33 wherein the compound is selected from
the compounds, their tautomers and their pharmaceutically
acceptable salts, of the group consisting of
4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridine;
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine;
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]methanesulfonamide;
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylsulfamide-
; [4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]urea;
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]sulfamide;
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine;
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylurea;
4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine;
4-[4-(4-fluorophenyl)-1-m- ethyl-1H-pyrazol-3-yl]pyridine;
4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyra- zole-1-ethanol;
4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazol-
e-1-ethanamine;
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]e-
thyl]morpholine; 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine;
1-methyl-4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]]piperi-
dine; and
1-methyl-4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-1-y-
l]piperidine.
42. The method of claim 34 wherein the disorder is a p38a kinase
mediated disorder.
43. The method of claim 34 wherein the P38 kinase mediated disorder
is selected from the group of disorders consisting of bone
resorption, graft vs. host reaction, atherosclerosis, arthritis,
osteoarthritis, rheumatoid arthritis, gout, psoriasis, topical
inflammatory disease state, adult respiratory distress syndrome,
asthma, chronic pulmonary inflammatory disease, cardiac reperfusion
injury, renal reperfusion injury, thrombus, glomerulonephritis,
Crohn's disease, ulcerative colitis, inflammatory bowel disease and
cachexia.
44. The method of claim 34 wherein the p38 kinase mediated disorder
is inflammation.
45. The method of claim 34 wherein the p38 kinase mediated disorder
is arthritis.
46. The method of claim 34 wherein the p38 kinase mediated disorder
is asthma.
47. The method of claim 34 wherein the compound is selected from
the compounds, their tautomers and their pharmaceutically
acceptable salts, of the group consisting of
4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridine;
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine;
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]methanesulfonamide;
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylsulfamide-
; [4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]urea;
[4-(4-fluorophenyl)-S-(4-pyridinyl)-1H-pyrazol-3-yl]sulfamide;
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine;
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylurea;
4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine;
4-(4-(4-fluorophenyl)-1-m- ethyl-1H-pyrazol-3-yl]pyridine;
4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyra- zole-1-ethanol;
4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazol-
e-1-ethanamine;
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]e-
thyl]morpholine; 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine;
1-methyl-4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]]piperi-
dine; and
1-methyl-4-[2-[4-(4-fluorophenyl)-S-(4-pyridinyl)-1H-pyrazol-1-y-
l]piperidine.
48. A compound of claim 1 that is
4-(4-(4-fluorophenyl)-1H-pyrazol-3-yl]py- ridine or a
pharmaceutically-acceptable salt or a tautomer thereof.
49. The method of claim 33 wherein the compound is
4-[4-(4-fluorophenyl)-1- H-pyrazol-3-yl]pyridine or a
pharmaceutically-acceptable salt or a tautomer thereof.
50. The method of claim 34 wherein the compound is
4-[4-(4-fluorophenyl)-1- H-pyrazol-3-yl]pyridine or a
pharmaceutically-acceptable salt or a tautomer thereof.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application claims priority from U.S. Provisional
Application Serial No. 60/047,535 filed May 22, 1997.
FIELD OF THE INVENTION
[0002] This invention relates to a novel group of pyrazole
compounds, compositions and methods for treating p38 kinase
mediated disorders.
BACKGROUND OF THE INVENTION
[0003] Mitogen-activated protein kinases (MAP) is a family of
proline-directed serine/threonine kinases that activate their
substrates by dual phosphorylation. The kinases are activated by a
variety of signals including nutritional and osmotic stress, UV
light, growth factors, endotoxin and inflammatory cytokines. The
p38 MAP kinase group is a MAP family of various isoforms, including
p38.alpha., p38.beta. and p38.gamma., and is responsible for
phosphorylating and activating transcription factors (e.g. ATF2,
CHOP and MEF2C) as well as other kinases (e.g. MAPKAP-2 and
MAPKAP-3). The p38 isoforms are activated by bacterial
lipopolysaccharide, physical and chemical stress and by
pro-inflammatory cytokines, including tumor necrosis factor
(TNF-.alpha.) and interleukin-1 (IL-1). The products of the p38
phosphorylation mediate the production of inflammatory cytokines,
including TNF and IL-1, and cyclooxygenase-2.
[0004] TNF-.alpha. is a cytokine produced primarily by activated
monocytes and macrophages. Excessive or unregulated TNF production
has been implicated in mediating a number of diseases. Recent
studies indicate that TNF has a causative role in the pathogenesis
of rheumatoid arthritis. Additional studies demonstrate that
inhibition of TNF has broad application in the treatment of
inflammation, inflammatory bowel disease, multiple sclerosis and
asthma.
[0005] TNF has also been implicated in viral infections, such as
HIV, influenza virus, and herpes virus including herpes simplex
virus type-1 (HSV-1), herpes simplex virus type-2 (HSV-2),
cytomegalovirus (CMV), varicella-zoster virus (VZV), Epstein-Barr
virus, human herpesvirus-6 (HHV-6), human herpesvirus-7 (HHV-7),
human herpesvirus-8 (HHV-8), pseudorabies and rhinotracheitis,
among others.
[0006] IL-8 is another pro-inflammatory cytokine, which is produced
by mononuclear cells, fibroblasts, endothelial cells, and
keratinocytes, and is associated with conditions including
inflammation.
[0007] IL-1 is produced by activated monocytes and macrochages and
is involved in the inflammatory response. IL-1 plays a role in many
pathophysiological responses including rheumatoid arthritis, fever
and reduction of bone resorption.
[0008] TNF, IL-1 and IL-8 affect a wide variety of cells and
tissues and are important inflammatory mediators of a wide variety
of disease states and conditions. The inhibition of these cytokines
by inhibition of the p38 kinase is of benefit in controlling,
reducing and alleviating many of these disease states.
[0009] Various pyrazoles have previously been described. U.S. Pat.
No. 4,000,281, to Beiler and Binon, describes 4,5-aryl, heteroaryl
substituted pyrazoles with antiviral activity against both RNA and
DNA viruses such as myxoviruses, adenoviruses, rhinoviruses, and
various viruses of the herpes group. WO 92/19615, published Nov.
12, 1992, describes pyrazoles as novel fungicides. U.S. Pat. No.
3,984,431, to Cueremy and Renault, describes derivatives of
pyrazole-5-acetic acid as having antiinflammatory activity.
Specifically, [1-isobutyl-3,4-diphenyl-- 1H-pyrazol-5-yl]acetic
acid is described. U.S. Pat. No. 3,245,093 to Hinsgen et al,
describes a process for preparing pyrazoles. WO 83/00330, published
Feb. 3, 1983, describes new process for the preparation of
diphenyl-3,4-methyl-5-pyrazole derivatives. WO 95/06036, published
for preparing pyrazole and its derivatives. U.S. Pat. No.
5,589,439, to T. Goto, et al., describes tetrazole derivatives and
their use as herbicides. EP 515041 describes pyrimidyl substituted
pyrazole derivatives as novel agricultural fungicides. Japanese
Patent 4,145,081 describes pyrazolecarboxylic acid derivatives as
herbicides used in paddy fields, dry fields as well as
non-agricultural areas. Japanese Patent 5,345,772 describes novel
pyrazole derivatives having potent inhibitory activity against
acetylcholinesterase.
[0010] Pyrazoles have been described for use in the treatment of
inflammation. Japanese Patent 5,017,470 describes synthesis of
pyrazole derivatives as anti-inflammatory, anti-rheumatic,
anti-bacterial and anti-viral drugs. EP 115640, published Dec. 30,
1983, describes 4-imidazolyl-pyrazole derivatives as inhibitors of
thromboxane synthesis.
3-(4-Isopropyl-1-methylcyclohex-1-yl)-4-(imidazol-1-yl)-1H-pyrazole
is specifically described. WO 97/01551, published Jan. 16, 1997,
describes pyrazole compounds as adenosine antagonists.
4-(3-Oxo-2,3-dihydropyridazi- n-6-yl)-3-phenylpyrazole is
specifically described. U.S. Pat. No. 5,134,142, to Matsuo et al.
describes 1,5-diaryl pyrazoles as having anti-inflammatory
activity.
[0011] U.S. Pat. No. 5,559,137 to Adams et al, describes novel
pyrazoles (1,3,4,-substituted) as inhibitors of cytokines used in
the treatment of cytokine diseases. Specifically,
3-(4-fluorophenyl)-1-(4-methylsulfinylph-
enyl)-4-(4-pyridyl)-5H-pyrazole is described. WO 96/03385,
published Feb. 8, 1996, describes 3,4-substituted pyrazoles, as
having anti-inflammatory activity. Specifically,
4-[(1-ethyl-4-(4-pyridyl)-5-trifluoromethyl-1H-py-
razol-3-yl]benzenesulfonamide is described.
[0012] The invention's pyrazolyl compounds are found to show
usefulness as p38 kinase inhibitors.
DESCRIPTION OF THE INVENTION
[0013] A class of substituted pyrazolyl compounds useful in
treating p38 mediated disorders is defined by Formula I: 2
[0014] wherein
[0015] R.sup.1 is selected from hydrido, alkyl, cycloalkyl,
alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene, haloalkyl,
hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthioalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, her erocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkylene; and
[0016] R.sup.2 is selected from hydrido, alkyl, alkenyl, alkynyl,
heterocyclyl, haloalkyl, heterocyclylalkyl, amino, alkylamino,
aminoalkyl, alkoxy, alkylthio, carboxy, alkoxycarbonyl,
carboxyalkyl, aminocarbonylamino, alkylaminocarbonylamino,
alkylsulfonyl, aminosulfonyl, alkylsulfonylamino,
aminosulfonylamino, alkylaminosulfonylamino, and alkynylamino;
wherein the heterocyclyl and heterocyclylalkyl groups are
optionally substituted with one or more radicals independently
selected from alkylthio, alkylsulfonyl, alkylsulfinyl, halo, alkyl,
alkoxy, aryloxy, aralkoxy, heterocyclyl, haloalkyl, amino, cyano,
and hydroxy; and
[0017] Ar.sup.1 is aryl optionally substituted with one or more
radicals independently selected from halo, alkyl, alkenyl, alkynyl,
alkoxy, alkenoxy, alkyldioxy, alkylthio, alkylsulfinyl,
alkylsulfonyl, amino, aminocarbonyl, cyano, alkoxycarbonyl, formyl,
aminosulfonyl, alkylamino, nitro, arylamino, alkylcarbonylamino,
halosulfonyl, aminoalkyl, and haloalkyl; and
[0018] HetAr.sup.2 is pyridinyl, pyrimidinyl or quinolinyl
optionally substituted with one or more radicals independently
selected from alkylthio, alkylsulfonyl, alkylsulfinyl, halo, alkyl,
heterocyclyl, alkoxy, aralkoxy, haloalkyl, amino, cyano, aralkyl,
alkylamino, cycloalkylamino, cycloalkenylamino, arylamino,
alkynylamino, and aralkylamino; or
[0019] a pharmaceutically-acceptable salt or a tautomer
thereof.
[0020] Compounds of Formula I would be useful for, but not limited
to, the treatment of any disorder or disease state in a human, or
other mammal, which is excacerbated or caused by excessive or
unregulated TNF or p38 kinase production by such mammal.
Accordingly, the present invention provides a method of treating a
cytokine-mediated disease which comprises administering an
effective cytokine-interfering amount of a compound of Formula I,
or a pharmaceutically acceptable salt or tautomer thereof.
[0021] Compounds of Formula I would be useful for, but not limited
to, the treatment of inflammation in a subject, and for use as
antipyretics for the treatment of fever. Compounds of the invention
would be useful to treat arthritis, including but not limited to,
rheumatoid arthritis, spondyloarthropathies, gouty arthritis,
osteoarthritis, systemic lupus erythematosus and juvenile
arthritis, osteoarthritis, gouty arthritis and other arthritic
conditions. Such compounds would be useful for the treatment of
pulmonary disorders or lung inflammation, including adult
respiratory distress syndrome, pulmonary sarcoidosis, asthma,
silicosis, and chronic pulmonary inflammatory disease. The
compounds are also useful for the treatment of viral and bacterial
infections, including sepsis, septic shock, gram negative sepsis,
malaria, meningitis, cachexia secondary to infection or malignancy,
cachexia secondary to acquired immune deficiency syndrome (AIDS),
AIDS, ARC (AIDS related complex), pneumonia, and herpesvirus. The
compounds are also useful for the treatment of bone resorption
diseases, such as osteoporosis, endotoxic shock, toxic shock
syndrome, reperfusion injury, autoimmune disease including graft
vs. host reaction and allograft rejections, cardiovascular diseases
including atherosclerosis, thrombosis, congestive heart failure,
and cardiac reperfusion injury, renal reperfusion injury, liver
disease and nephritis, and myalgias due to infection.
[0022] The compounds are also useful for the treatment of
influenza, multiple sclerosis, cancer, diabetes, systemic lupus
erthrematosis (SLE), skin-related conditions such as psoriasis,
eczema, burns, dermatitis, keloid formation, and scar tissue
formation. Compounds of the invention also would be useful to treat
gastrointestinal conditions such as inflammatory bowel disease,
Crohn's disease, gastritis, irritable bowel syndrome and ulcerative
colitis. The compounds would also be useful in the treatment of
ophthalmic diseases, such as retinitis, retinopathies, uveitis,
ocular photophobia, and of acute injury to the eye tissue.
Compounds of the invention also would be useful for treatment of
angiogenesis, including neoplasia; metastasis; ophthalmological
conditions such as corneal graft rejection, ocular
neovascularization, retinal neovascularization including
neovascularization following injury or infection, diabetic
retinopathy, retrolental fibroplasia and neovascular glaucoma;
ulcerative diseases such as gastric ulcer; pathological, but
non-malignant, conditions such as hemaginomas, including invantile
hemaginomas, angiofibroma of the nasopharynx and avascular necrosis
of bone; diabetic nephropathy and cardiomyopathy; and disorders of
the female reproductive system such as endometriosis. The compounds
of the invention may also be useful for preventing the production
of cyclooxygenase-2.
[0023] Besides being useful for human treatment, these compounds
are also useful for veterinary treatment of companion animals,
exotic animals and farm animals, including mammals, rodents, and
the like. More preferred animals include horses, dogs, and
cats.
[0024] The present compounds may also be used in co-therapies,
partially or completely, in place of other conventional
antiinflammatories, such as together with steroids,
cyclooxygenase-2 inhibitors, NSAIDs, DMARDS, immunosuppressive
agents, 5-lipoxygenase inhibitors, LTB.sub.4 antagonists and
LTA.sub.4 hydrolase inhibitors.
[0025] As used herein, the term "TNF mediated disorder" refers to
any and all disorders and disease states in which TNF plays a role,
either by control of TNF itself, or by TNF causing another monokine
to be released, such as but not limited to IL-1, IL-6 or IL-8. A
disease state in which, for instance, IL-1 is a major component,
and whose production or action, is exacerbated or secreted in
response to TNF, would therefore be considered a disorder mediated
by TNF.
[0026] As used herein, the term "p38 mediated disorder" refers to
any and all disorders and disease states in which p38 plays a role,
either by control of p38 itself, or by p38 causing another factor
to be released, such as but not limited to IL-1, IL-6 or IL-8. A
disease state in which, for instance, IL-1 is a major component,
and whose production or action, is exacerbated or secreted in
response to p38, would therefore be considered a disorder mediated
by p38.
[0027] As TNF-.beta. has close structural homology with TNF-.alpha.
(also known as cachectin), and since each induces similar biologic
responses and binds to the same cellular receptor, the synthesis of
both TNF-.alpha. and TNF-.beta. are inhibited by the compounds of
the present invention and thus are herein referred to collectively
as "TNF" unless specifically delineated otherwise
[0028] A preferred class of compounds consists of those compounds
of Formula I wherein
[0029] R.sup.1 is selected from hydrido, lower alkyl, lower
cycloalkyl, lower cycloalkylalkylene, lower haloalkyl, lower
hydroxyalkyl, lower alkenyl, lower alkynyl, lower heterocyclyl,
lower aralkyl, lower alkoxyalkyl, lower mercaptoalkyl, lower
alkylthioalkylene, amino, lower alkylamino, lower arylamino, lower
aminoalkyl, lower alkylaminoalkylene, lower heterocyclylalkylene,
lower aminocarbonylalkylene, and lower alkylaminocarbonylalkylene;
and
[0030] R.sup.2 is selected from hydrido, lower alkyl, lower
alkenyl, lower alkynyl, lower haloalkyl, lower heterocyclyl, lower
heterocyclylalkylene, amino, lower alkylamino, lower alkynylamino,
lower aminoalkyl, lower alkylthio, lower carboxy, lower
alkoxycarbonyl, lower carboxyalkyl, lower aminocarbonylamino, lower
alkylaminocarbonylamino, lower alkylsulfonyl, lower aminosulfonyl,
lower alkylsulfonylamino, lower aminosulfonylamino, and lower
alkylaminosulfonylamino, wherein the heterocyclyl and
heterocyclylalkyl groups are optionally substituted with one or
more radicals independently selected from lower alkylthio, lower
alkylsulfonyl, lower alkylsulfinyl, halo, lower alkyl, lower
alkoxy, aryloxy, lower heterocyclyl, lower haloalkyl, amino, and
cyano; and
[0031] Ar.sup.1 is selected from phenyl, biphenyl, and naphthyl,
wherein Ar.sup.1 is optionally substituted with one or more
radicals independently selected from lower alkylthio, lower
alkylsulfonyl, aminosulfonyl, halo, lower alkyl, lower alkenyl,
lower alkynyl, lower alkylsulfinyl, cyano, lower alkoxycarbonyl,
aminocarbonyl, formyl, lower alkylcarbonylamino, lower haloalkyl,
lower alkoxy, lower alkenyloxy, lower alkyldioxy, amino, lower
alkylamino, lower aminoalkyl, arylamino, nitro, and halosulfonyl;
and
[0032] HetAr.sup.2 is pyridinyl or pyrimidinyl optionally
substituted with one or more radicals independently selected from
lower alkylthio, lower alkylsulfonyl, lower alkylsulfinyl, halo,
lower alkyl, lower heterocyclyl, lower alkoxy, lower aralkoxy,
lower haloalkyl, amino, cyano, lower aralkyl, lower alkylamino,
lower cycloalkylamino, lower arylamino, lower alkynylamino, and
lower aralkylamino; or
[0033] a pharmaceutically-acceptable salt or tautomer thereof.
[0034] A class of compounds of particularly interest consists of
these compounds of Formula I wherein
[0035] R.sup.1 is selected from hydrido, methyl, ethyl, isopropyl,
tert-butyl, isobutyl, trichloroethyl, pentafluoroethyl,
heptafluoropropyl, difluoroethyl, difluoropropyl, dichloroethyl,
dichloropropyl, vinyl, allyl, ethynyl, propargyl, morpholinyl,
piperidinyl, piperazinyl, benzyl, phenylethyl, morpholinomethyl,
morpholinoethyl, pyrrolidinylmethyl, piperazinylmethyl,
piperidinylmethyl, pyridinylmethyl, thienylmethyl, methoxymethyl,
ethoxymethyl, amino, methylamino, dimethylamino, phenylamino,
methylaminomethyl, dimethylaminomethyl, methylaminoethyl,
dimethylaminoethyl, cyclopropyl, cyclopentyl, cyclohexyl,
cyclohexylmethyl, hydroxymethyl, hydroxyethyl, methylthio, and
methylthiomethyl; and
[0036] R.sup.2 is selected from hydrido, methyl, ethyl, propyl,
isopropyl, tert-butyl, isobutyl, fluoromethyl, difluoromethyl,
trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl,
pentafluoroethyl, heptafluoropropyl, difluorochloromethyl,
dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl,
dichloropropyl, amino, N-methylamino, N,N-dimethylamino,
ethynylamino, propargylamino, piperidinyl, piperazinyl,
morpholinomethyl, pyrrolidinylmethyl, piperazinylmethyl,
piperidinylmethyl, pyridinylmethyl, thienylmethyl, thiazolylmethyl,
oxazolylmethyl, pyrimidinylmethyl, quinolylmethyl,
isoquinolinylmethyl, imidazolylmethyl, benzimidazolylmethyl,
furylmethyl, pyrazinylmethyl, aminocarbonylamino,
methylaminocarbonylamino, dimethylaminocarbonylamino,
ethylaminocarbonylamino, diethylaminocarbonylamino,
methylsulfonylamino, ethylsulfonylamino, aminosulfonylamino,
methylaminosulfonylamino, dimethylaminosulfonylamino,
ethylaminosulfonylamino, and diethylaminosulfonylamino; and
[0037] Ar.sup.1 is selected from phenyl, biphenyl, and naphthyl,
wherein Ar.sup.1 is optionally substituted with one or more
radicals independently selected from methylthio, methylsulfinyl,
methylsulfonyl, fluoro, chloro, bromo, aminosulfonyl, methyl,
ethyl, isopropyl, tert-butyl, isobutyl, cyano, methoxycarbonyl,
ethoxycarbonyl, aminocarbonyl, methylcarbonylamino,
trifluoromethyl, difluoromethyl, fluoromethyl, trichloromethyl,
dichloromethyl, chloromethyl, allyl, vinyl, ethynyl, propargyl,
methoxy, ethoxy, propyloxy, n-butoxy, amino, methylamino,
ethylamino, dimethylamino, diethylamino, aminomethyl, aminoethyl,
N-methyl, N-phenylamino, phenylamino, diphenylamino, nitro, and
chlorosulfonyl; and
[0038] HetAr.sup.2 is selected from pyridinyl and pyrimidinyl,
wherein HetAr.sup.2 is optionally substituted with one or more
radicals independently selected from methylthio, methylsulfinyl,
methylsulfonyl, fluoro, chloro, bromo, methyl, ethyl, isocropyl,
tert-butyl, isobutyl, methoxyl, ethoxyl, phenoxyl, benzoxyl,
phenethyl, trifluoromerhyl, flucromethyl, difluoromethyl, amino,
benzylamino, propargylamino, cyclopropylamino, cyclobutylamino,
cyclopentylamino, and cyano; or
[0039] a pharmaceutically-acceptable salt or tautomer thereof.
[0040] A class of compounds of specific interest consists of those
compounds of Formula I wherein
[0041] R.sup.1 is hydrido, methyl, ethyl, hydroxyethyl, propargyl,
dimethylaminoethyl or morpholinoethyl; and
[0042] R.sup.2 is selected from hydrido, methyl, ethyl, amino,
aminocarbonylamino, methylaminocarbonylamino, methylsulfonylamino,
aminosulfonylamino, and methylaminosulfonylamino; and
[0043] Ar.sup.1 is phenyl optionally substituted with one or more
radicals independently selected from methylthio, methylsulfinyl,
methylsulfonyl, fluoro, chloro, bromo, aminosulfonyl, methyl,
ethyl, isopropyl, tert-butyl, isobutyl, cyano, methoxycarbonyl,
ethoxycarbonyl, aminocarbonyl, methylcarbonylamino,
trifluoromethyl, difluoromethyl, fluoromethyl, trichloromethyl,
dichloromethyl, chloromethyl, methoxy, ethoxy, propyloxy, n-butoxy,
amino, methylamino, ethylamino, dimethylamino, diethylamino,
aminomethyl, aminoethyl, N-methyl, N-phenylamino, phenylamino,
diphenylamino, nitro, and chlorosulfonyl; and
[0044] HetAr.sup.2 is optionally substituted with one or more
radicals independently selected from methylthio, methylsulfinyl,
methylsulfonyl, fluoro, chloro, bromo, methyl, ethyl, isopropyl,
tert-butyl, isobutyl, methoxyl, ethoxyl, phenoxyl, benzoxyl,
trifluoromethyl, fluoromethyl, difluoromethyl, amino,
propargylamino, and cyano; or
[0045] a pharmaceutically-acceptable salt or a tautomer
thereof.
[0046] A class of compounds of very specific interest consists of
those compounds of Formula I wherein
[0047] R.sup.1 is hydrido or methyl; and
[0048] R.sup.2 is hydrido or methyl; and
[0049] Ar.sup.1 is phenyl which is optionally substituted with one
or more radicals independently selected fluoro, chloro, methyl,
ethyl, trifluoromethyl, methoxy, ethoxy, dimethylamino, and nitro;
and
[0050] HetAr.sup.2 is optionally substituted with one or more
radicals independently selected from methyl, chloro, fluoro, and
trifluoromethyl; or
[0051] a pharmaceutically-acceptable salt or tautomer thereof.
[0052] A family of specific compounds of particular interest within
Formula I consists of compounds, and tautomers and
pharmaceutically-acceptable salts thereof, as follows:
[0053] 4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridine;
[0054] 4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine;
[0055]
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]methanesulfona-
mide;
[0056]
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylsul-
famide;
[0057]
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]urea;
[0058]
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]sulfamide;
[0059]
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine;
[0060]
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylure-
a;
[0061] 4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine;
[0062] 4-[4-(4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine;
[0063]
4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazole-1-ethanol;
[0064]
4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazole-1-ethan-
amine;
[0065]
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]ethyl]morp-
holine;
[0066] 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine;
[0067] 4-(4-phenyl-1H-pyrazol-5-yl)pyridine;
[0068]
1-methyl-4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]]-
piperidine; and
[0069]
1-methyl-4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-1-yl]p-
iperidine.
[0070] The term "hydrido" denotes a single hydrogen atom (H). This
hydrido radical may be attached, for example, to an oxygen atom to
form a hydroxyl radical or two hydrido radicals may be attached to
a carbon atom to form a methylene (--CH.sub.2--) radical. Where
used, either alone or within other terms such as "haloalkyl",
"alkylsulfonyl", "alkoxyalkyl", "hydroxyalkyl", "mercaptoalkyl",
the term "alkyl" embraces linear or branched radicals having one to
about twenty carbon atoms or, preferably, one to about twelve
carbon atoms. More preferred alkyl radicals are "lower alkyl"
radicals having one to about ten carbon atoms. Most preferred are
lower alkyl radicals having one to about six carbon atoms. Examples
of such radicals include methyl, ethyl, n-propyl, isopropyl,
n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl
and the like. The term "alkenyl" embraces linear or branched
radicals having at least one carbon-carbon double bond of two to
about twenty carbon atoms or, preferably, two to about twelve
carbon atoms. More preferred alkenyl radicals are "lower alkenyl"
radicals having two to about six carbon atoms. Examples of alkenyl
radicals include ethenyl, propenyl, allyl, propenyl, butenyl and
4-methylbutenyl. The terms "alkenyl" and "lower alkenyl", embrace
radicals having "cis" and "trans" orientations, or alternatively,
"E" and "Z" orientations. The term "alkynyl" embraces linear or
branched radicals having at least one carbon-carbon triple bond of
two to about twenty carbon atoms or, preferably, two to about
twelve carbon atoms. More preferred alkynyl radicals are "lower
alkynyl" radicals having two to about six carbon atoms. Examples of
alkynyl radicals include propargyl, 1-propynyl, 2-propynyl,
1-butyne, 2-butenyl and 1-pentynyl. The term "cycloalkyl" embraces
saturated carbocyclic radicals having three to about twelve carbon
atoms. More preferred cycloalkyl radicals are "lower cycloalkyl"
radicals having three to about eight carbon atoms. Examples of such
radicals include cyclopropyl, cyclobutyl, cyclopentyl and
cyclohexyl. The term "cycloalkylalkylene" embraces alkyl radicals
substituted with a cycloalkyl radical. More preferred
cycloalkylalkylene radicals are "lower cycloalkylalkylene" which
embrace lower alkyl radicals substituted with a lower cycloalkyl
radical as defined above. Examples of such radicals include
cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl and
cyclohexylmethyl. The term "cycloalkenyl" embraces partially
unsaturated carbocyclic radicals having three to twelve carbon
atoms and one or two double bonds but not necessarily conjugated
("cycloalkyldienyl"). More preferred cycloalkenyl radicals are
"lower cycloalkenyl" radicals having four to about eight carbon
atoms. Examples of such radicals include cyclobutenyl,
cyclopentenyl and cyclohexenyl. The term "cycloalkenylalkylene"
embraces alkyl radicals substituted with a cycloalkenyl radical.
More preferred cycloalkenylalkylene radicals are "lower
cycloalkenylalkylene" which embrace lower alkyl radicals
substituted with a lower cycloalkenyl radical, as defined above.
Examples of such radicals include cyclobutenylmethyl,
cyclopentenylmethyl and cyclohexenylmethyl. The term "halo" means
halogens such as fluorine, chlorine, bromine or iodine. The term
"haloalkyl" embraces radicals wherein any one or more of the alkyl
carbon atoms is substituted with halo as defined above.
Specifically embraced are monohaloalkyl, dihaloalkyl and
polyhaloalkyl radicals. A monohaloalkyl radical, for one example,
may have either an iodo, bromo, chloro or fluoro atom within the
radical. Dihalo and polyhaloalkyl radicals may have two or more of
the same halo atoms or a combination of different halo radicals.
"Lower haloalkyl" embraces radicals having one to six carbon atoms.
Examples of haloalkyl radicals include fluoromethyl,
difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl,
trichloromethyl, pentafluoroethyl, heptafluoropropyl,
difluorochloromethyl, dichlorofluoromethyl, difluoroethyl,
difluoropropyl, dichloroethyl and dichloropropyl. The term
"hydroxyalkyl" embraces linear or branched alkyl radicals having
one to about ten carbon atoms, any one of which may be substituted
with one or more hydroxyl radicals. More preferred hydroxyalkyl
radicals are "lower hydroxyalkyl" radicals having one to six carbon
atoms and one or more hydroxyl radicals. Examples of such radicals
include hydroxymethyl, hydroxyethyl, hydroxypropyl, hydroxybutyl
and hydroxyhexyl. The terms "alkoxy" and "alkyloxy" embrace linear
or branched oxy-containing radicals each having alkyl portions of
one to about ten carbon atoms. More preferred alkoxy radicals are
"lower alkoxy" radicals having one to six carbon atoms. Examples of
such radicals include methoxy, ethoxy, propoxy, butoxy and
tert-butoxy. The term "alkoxyalkyl" embraces alkyl radicals having
one or more alkoxy radicals attached to the alkyl radical to form,
for example, monoalkoxyalkyl and dialkoxyalkyl radicals. The
"alkoxy" radicals may be further substituted with one or more halo
atoms, such as fluoro, chloro or bromo, to provide "haloalkoxy"
radicals.
[0071] The term "aryl", alone or in combination, means a
carbocyclic aromatic system containing one, two or three rings
wherein such rings may be attached together in a pendent manner or
may be fused. More preferred aryl are 6-12 membered aryl radicals.
Examples of such radicals include phenyl, naphthyl,
tetrahydronaphthyl, indane and biphenyl. Aryl moieties may also be
substituted at a substitutable position with one or more
substituents selected independently from, for example, halo, alkyl,
alkenyl, alkynyl, alkoxy, alkenoxy, alkyldioxy, alkylthio,
alkylsulfinyl, alkylsulfonyl, amino, aminocarbonyl, cyano,
alkoxycarbonyl, formyl, aminosulfonyl, alkylamino, nitro,
arylamino, alkylcarbonylamino, halosulfonyl, aminoalkyl, and
haloalkyl, alkoxyalkyl, alkylaminoalkyl, carboxyalkyl,
alkoxycarbonylalkyl, aminocarbonylalkyl, aralkoxy, hydroxyl, acyl,
carboxy, aminocarbonyl, and aralkoxycarbonyl. The term
"alkyldioxyl" encompasses an alkyldioxy bridge, such as a
methylenedioxy bridge, between two carbon ring atoms of an aryl
moiety.
[0072] The term "heterocyclyl" embraces saturated, partially
unsaturated and unsaturated heteroatom-containing ring-shaped
radicals, which can also be called "heterocyclyl",
"heterocycloalkenyl" and "heteroaryl" correspondingly, where the
heteroatoms may be selected from nitrogen, sulfur and oxygen.
Examples of saturated heterocyclyl radicals include saturated 3 to
6-membered heteromonocyclic group containing 1 to 4 nitrogen atoms
(e.g. pyrrolidinyl, imidazolidinyl, piperidino, piperazinyl, etc.);
saturated 3 to 6-membered heteromonocyclic group containing 1 to 2
oxygen atoms and 1 to 3 nitrogen atoms (e.g. morpholinyl, etc.);
saturated 3 to 6-membered heteromonocyclic group containing 1 to 2
sulfur atoms and 1 to 3 nitrogen atoms (e.g., thiazolidinyl, etc.).
Examples of partially unsaturated heterocyclyl radicals include
dihydrothiophene, dihydropyran, dihydrofuran and dihydrothiazole.
Heterocyclyl radicals may include a pentavalent nitrogen, such as
in tetrazolium and pyridinium radicals. The term "heteroaryl"
embraces unsaturated heterocyclyl radicals. Examples of heteroaryl
radicals include unsaturated 3 to 6 membered heteromonocyclic group
containing 1 to 4 nitrogen atoms, for example, pyrrolyl,
pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidyl, pyrazinyl,
pyridazinyl, triazolyl (e.g., 4H-1,2,4-triazolyl,
1H-1,2,3-triazolyl, 2H-1,2,3-triazolyl, etc.) tetrazolyl (e.g.
1H-tetrazolyl, 2H-tetrazolyl, etc.), etc.; unsaturated condensed
heterocyclyl group containing 1 to 5 nitrogen atoms, for example,
indolyl, isoindolyl, indolizinyl, benzimidazolyl, quinolyl,
isoquinolyl, indazolyl, benzotriazolyl, tetrazolopyridazinyl (e.g.,
tetrazolo[1,5-b]pyridazinyl, etc.), etc.; unsaturated 3 to
6-membered heteromonocyclic group containing an oxygen atom, for
example, pyranyl, furyl, etc.; unsaturated 3 to 6-membered
heteromonocyclic group containing a sulfur atom, for example,
thienyl, etc.; unsaturated 3- to 6-membered heteromonocyclic group
containing 1 to 2 oxygen atoms and 1 to 3 nitrogen atoms, for
example, oxazolyl, isoxazolyl, oxadiazolyl (e.g.,
1,2,4-oxadiazolyl, 1,3,4-oxadiazolyl, 1,2,5-oxadiazolyl, etc.)
etc.; unsaturated condensed hecerocyclyl group containing 1 to 2
oxygen atoms and 1 to 3 nitrogen atoms (e.g. benzoxazolyl,
benzoxadiazolyl, etc.); unsaturated 3 to 6-membered
heteromonocyclic group containing 1 to 2 sulfur atoms and 1 to 3
nitrogen atoms, for example, thiazolyl, thiadiazolyl (e.g.,
1,2,4-thiadiazolyl, 1,3,4-thiadiazolyl, 1,2,5-thiadiazolyl, etc.)
etc.; unsaturated condensed heterocyclyl group containing 1 to 2
sulfur atoms and 1 to 3 nitrogen atoms (e.g., benzothiazolyl,
benzothiadiazclyl, etc.) and the like. The term "heteroaryl" also
embraces radicals where heterocyclyl radicals are fused with aryl
radicals. Examples of such fused bicyclic radicals include
benzofuran, benzothiophene, and the like. Said heterocyclyl group
may have 1 to 3 substituents such as alkyl, hydroxyl, halo, alkoxy,
oxo, amino and alkylamino. The term "heterocyclylalkylene" embraces
heterocyclyl-substituted alkyl radicals. More preferred
heterocyclylalkylene radicals are "lower heterocyclylalkylene"
radicals having one to six carbon atoms and a heterocyclyl
radical.
[0073] The term "alkylthio" embraces radicals containing a linear
or branched alkyl radical, of one to about ten carbon atoms
attached to a divalent sulfur atom. More preferred alkylthio
radicals are "lower alkylthio" radicals having alkyl radicals of
one to six carbon atoms. Examples of such lower alkylthio radicals
are methylthio, ethylthio, propylthio, butylthio and hexylthio. The
term "alkylthioalkylene" embraces radicals containing an alkylthio
radical attached through the divalent sulfur atom to an alkyl
radical of one to about ten carbon atoms. More preferred
alkylthioalkylene radicals are "lower alkylthioalkylene" radicals
having alkyl radicals of one to six carbon atoms. Examples of such
lower alkylthioalkylene radicals include methylthiomethyl. The term
"alkylsulfinyl" embraces radicals containing a linear or branched
alkyl radical, of one to about ten carbon atoms, attached to a
divalent --S(.dbd.O)-- radical. More preferred alkylsulfinyl
radicals are "lower alkylsulfinyl", radicals having alkyl radicals
of one to six carbon atoms. Examples of such lower alkylsulfinyl
radicals include methylsulfinyl, ethylsulfinyl, butylsulfinyl and
hexylsulfinyl. The term "sulfonyl", whether used alone or linked to
other terms such as "alkylsulfonyl", or "halosulfonyl" denotes a
divalent radical, --SO.sub.2--. "Alkylsulfonyl" embraces alkyl
radicals attached to a sulfonyl radical, where alkyl is defined as
above. More preferred alkylsulfonyl radicals are "lower
alkylsulfonyl" radicals having one to six carbon atoms. Examples of
such lower alkylsulfonyl radicals include methylsulfonyl,
ethylsulfonyl and propylsulfonyl. The "alkylsulfonyl" radicals may
be further substituted with one or more halo atoms, such as fluoro,
chloro or bromo, to provide haloalkylsulfonyl radicals. The term
"halosulfonyl" embraces halo radicals attached to a sulfonyl
radical. Examples of such halosulfonyl radicals include
chlorosulfonyl and bromosulfonyl. The terms "sulfamyl",
"aminosulfonyl" and "sulfonamidyl" denote NH.sub.2O.sub.2S--.
[0074] The term "carbonyl", whether used alone or with other terms,
such as "alkoxycarbonyl", denotes --(C.dbd.O)--. The terms
"carboxy" or "carboxyl", whether used alone or with other terms,
such as "carboxyalkyl", denotes --CO.sub.2H. The term
"carboxyalkyl" embraces alkyl radicals substituted with a carboxy
radical. More preferred are "lower carboxyalkyl" radicals which
embrace carboxy-substituted lower alkyl radicals, as defined above.
Examples of such lower carboxyalkyl radicals include carboxymethyl,
carboxyethyl and carboxypropyl. The term "alkoxycarbonyl" means a
radical containing an alkoxy radical, as defined above, attached
via an oxygen atom to a carbonyl radical. More preferred are "lower
alkoxycarbonyl" radicals with alkyl portions having one to six
carbons. Examples of such lower alkoxycarbonyl (ester) radicals
include methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl,
butoxycarbonyl and hexyloxycarbonyl. The term
"alkoxycarbonylalkylene" embraces alkyl radicals substituted with
an alkoxycarbonyl radical as defined above. More preferred are
"lower alkoxycarbonylalkylene" radicals with alkyl portions having
one to six carbons. Examples of such lower alkoxycarbonylalkylene
radicals include methoxycarbonylmethylene, ethoxycarbonylmethylene,
methoxycarbonylethylene and ethoxycarbonylethylene. The term
"alkylcarbonyl", includes radicals having alkyl radicals attached
to a carbonyl radical. Examples of such radicals include
methylcarbonyl, ethylcarbonyl, propylcarbonyl, butylcarbonyl, and
pentylcarbonyl. The term "aralkyl" embraces aryl-substituted alkyl
radicals. Preferred aralkyl radicals are "lower aralkyl", having
lower alkyl groups substituted with one or more aryl groups.
Examples of such groups include benzyl, diphenylmethyl,
triphenylmethyl, phenylethyl, and diphenylethyl. The aryl in said
aralkyl may be additionally substituted with halo, alkyl, alkoxy,
haloalkyl and haloalkoxy. The terms benzyl and phenylmethyl are
interchangeable. The term "heterocyclylalkylene" embraces
saturated, partially unsaturated and unsaturated
heterocyclyl-substituted alkyl radicals such as pyrrolidinylmethyl,
pyridylmethyl, quinolylmethyl, thienylmethyl, furylethyl, and
quinolylethyl. The heteroaryl in heteroaralkyl (unsaturated
heterocyclyl-substituted alkyl radicals) may be additionally
substituted with halo, alkyl, alkoxy, haloalkyl and haloalkoxy. The
term "aryloxy" embraces aryl radicals attached through an oxygen
atom to other radicals. The term "aralkoxy" embraces aralkyl
radicals attached through an oxygen atom to other radicals.
[0075] The term "aminoalkyl" embraces alkyl radicals substituted
with amino radicals. More preferred are "lower aminoalkyl"
radicals. Examples of such radicals include aminomethyl,
aminoethyl, and the like. The term "alkylamino" denotes amino
groups which are substituted with one or two alkyl radicals.
Preferred are "lower alkylamino" radicals having alkyl portions
having one to six carbon atoms. Suitable lower alkylamino may be
monosubstituted N-alkylamino or disubstituted N,N-alkylamino, such
as N-methylamino, N-ethylamino, N,N-dimethylamino, N,N-diethylamino
or the like. The term "arylamino" denotes amino groups which are
substituted with one or two aryl radicals, such as N-phenylamino.
The "arylamino" radicals may be further substituted on the aryl
ring portion of the radical. The term "aminocarbonyl" denotes an
amide group of the formula --C(.dbd.O)NH. The term
"alkylaminocarbonyl" denotes an aminocarbonyl group which has been
substituted with one or two alkyl radicals on the amino nitrogen
atom. Preferred are "N-alkylaminocarbonyl" and
"N,N-dialkylaminocarbonyl" radicals. More preferred are "lower
N-alkylaminocarbonyl" and "lower N,N-dialkylaminocarbonyl" radicals
with lower alkyl portions as defined above. The term
"aminocarbonylamino" embraces radicals having one or more
aminocarbonyl radicals attached to an amino radical. The term
"alkylaminocarbonylamino" embraces radicals having one or more
alkyl radicals attached to an aminocarbonylamino radical. Preferred
are "lower alkylaminocarbonylamino" radicals with lower alkyl
portions as defined above. The term "alkylcarbonylamino" embraces
amino groups which are substituted with one or more alkylcarbonyl
radicals. More preferred alkylcarbonylamino radicals are "lower
alkylcarbonylamino" having lower alkylcarbonyl radicals as defined
above attached to amino radicals. The term "alkylaminoalkylene"
embraces radicals having one or more alkyl radicals attached to an
aminoalkyl radical. The term "alkylsulfonylamino" embraces radicals
having one or more alkylsulfonyl radicals attached to an amino
radical. Preferred are "lower alkylsulfonylamino" radicals with
lower alkyl portions as defined above. The term
"aminosulfonylamino" embraces radicals having one or more
aminosulfonyl radicals attached to an amino radical. The term
"alkylaminosulfonylamino" embraces radicals having one or more
alkyl radicals attached to an aminosulfonylamino radical. Preferred
are "lower alkylaminosulfonylamino" radicals with lower alkyl
portions as defined above.
[0076] The additional terms used to describe the substituents of
the pyrazole ring and not specifically defined herein are defined
in a similar manner to that illustrated in the above definitions.
As above, more preferred substituents are those containing "lower"
radicals. Unless otherwise defined to contrary, the term "lower" as
used in this application means that each alkyl radical of a
pyrazole ring substituent comprising one or more alkyl radicals has
one to about six carbon atoms; each alkenyl radical of a pyrazole
ring substituent comprising one or more alkenyl radicals has two to
about six carbon atoms; each alkynyl radical of a pyrazole ring
substituent comprising one or more alkynyl radicals has two to
about six carbon atoms; each cycloalkyl or cycloalkenyl radical of
a pyrazole ring substituent comprising one or more cycloalkyl
and/or cycloalkenyl radicals is a 3 to 8 membered ring cycloalkyl
or cycloalkenyl radical, respectively; each aryl radical of a
pyrazole ring substituent comprising one or more aryl radicals is a
monocyclic aryl radical; and each heterocyclyl radical of a
pyrazole ring substituent comprising one or more heterocyclyl
radicals is a 4-8 membered ring heterocyclyl.
[0077] The present invention comprises the tautomeric forms of
compounds of formula I. As illustrated below, the pyrazoles of
Formula I and I' are magnetically and structurally equivalent
because of the prototropic tautomeric nature of the hydrogen: 3
[0078] The present invention also comprises compounds of Formula I
having one or more asymmetric carbons. It is known to those skilled
in the art that those pyrazoles of the present invention having
asymmetric carbon atoms may exist in diastereomeric, racemic, or
optically active forms. All of these forms are contemplated within
the scope of this invention. More specifically, the present
invention includes enantiomers, diastereomers, racemic mixtures,
and other mixtures thereof.
[0079] The present invention comprises a pharmaceutical composition
for the treatment of a TNF mediated disorder, a p38 kinase mediated
disorder, inflammation, and/or arthritis, comprising a
therapeutically-effective amount of a compound of Formula I, or a
therapeutically-acceptable salt or tautomer thereof, in association
with at least one pharmaceutically-acceptable carrier, adjuvant or
diluent.
[0080] The present invention also comprises a therapeutic method of
treating a TNF mediated disorder, a p38 kinase mediated disorder,
inflammation and/or arthritis in a subject, the method comprising
treating a subject having or susceptible to such disorder or
condition with a therapeutically-effective amount of a compound of
Formula 4
[0081] wherein
[0082] R.sup.1 is selected from hydrido, alkyl, cycloalkyl,
alkenyl, alkynyl, heterocyclyl, cycloalkylalkylene, haloalkyl,
hydroxyalkyl, aralkyl, alkoxyalkyl, mercaptoalkyl,
alkylthioalkylene, amino, alkylamino, arylamino, aminoalkyl,
alkylaminoalkylene, heterocyclylalkylene, aminocarbonylalkylene,
and alkylaminocarbonylalkyle- ne; and
[0083] R.sup.2 is selected from hydrido, alkyl, alkenyl, alkynyl,
heterocyclyl, haloalkyl, heterocyclylalkyl, amino, alkylamino,
aminoalkyl, alkoxy, alkylthio, carboxy, alkoxycarbonyl,
carboxyalkyl, aminocarbonylamino, alkylaminocarbonylamino,
alkylsulfonyl, aminosulfonyl, alkylsulfonylamino,
aminosulfonylamino, alkylaminosulfonylamino, and alkynylamino;
wherein the heterocyclyl and heterocyclylalkyl groups are
optionally substituted with one or more radicals independently
selected from alkylthio, alkylsulfonyl, alkylsulfinyl, halo, alkyl,
alkoxy, aryloxy, aralkoxy, heterocyclyl, haloalkyl, amino, cyano,
and hydroxy; and
[0084] Ar.sup.1 is aryl optionally substituted with one or more
radicals independently selected from halo, alkyl, alkenyl, alkynyl,
alkoxy, alkenoxy, alkyldioxy, alkylthio, alkylsulfinyl,
alkylsulfonyl, amino, aminocarbonyl, cyano, alkoxycarbonyl, formyl,
aminosulfonyl, alkylamino, nitro, arylamino, alkylcarbonylamino,
halosulfonyl, aminoalkyl, and haloalkyl; and
[0085] HetAr.sup.2 is pyridinyl, pyrimidinyl or quinolinyl
optionally substituted with one or more radicals independently
selected from alkylthio, alkylsulfonyl, alkylsulfinyl, halo, alkyl,
heterocyclyl, alkoxy, aralkoxy, haloalkyl, amino, cyano, aralkyl,
alkylamino, cycloalkylamino, cycloalkenylamino, arylamino,
alkynylamino, and aralkylamino; or a pharmaceutically-acceptable
salt or a tautomer thereof.
[0086] Also included in the family of compounds of Formula I are
the pharmaceutically-acceptable salts thereof. The term
"pharmaceutically-acceptable salts" embraces salts commonly used to
form alkali metal salts and to form addition salts of free acids or
free bases. The nature of the salt is not critical, provided that
it is pharmaceutically-acceptable. Suitable
pharmaceutically-acceptable acid addition salts of compounds of
Formula I may be prepared from an inorganic acid or from an organic
acid. Examples of such inorganic acids are hydrochloric,
hydrobromic, hydroiodic, nitric, carbonic, sulfuric and phosphoric
acid. Appropriate organic acids may be selected from aliphatic,
cycloaliphatic, aromatic, araliphatic, heterocyclyl, carboxylic and
sulfonic classes of organic acids, example of which are formic,
acetic, propionic, succinic, glycolic, gluconic, lactic, malic,
tartaric, citric, ascorbic, glucuronic, maleic, fumaric, pyruvic,
aspartic, glutamic, benzoic, anthranilic, mesylic, stearic,
salicylic, p-hydroxybenzoic, phenylacetic, mandelic, embonic
(pamoic), methanesulfonlc, ethanesulfonic, benzenesulfonic,
pantothenic, toluenesulfonic, 2-hydroxyethanesulfonic, sulfanilic,
cyclohexylaminosulfonic, algenic, .beta.-hydroxybutyric, galactaric
and galacturonic acid. Suitable pharmaceutically-acceptable base
addition salts of compounds of Formula I include metallic salts and
organic salts. More preferred metallic salts include, but are not
limited to appropriate alkali metal (group Ia) salts, alkaline
earth metal (group IIa) salts and other physiological acceptable
metals. Such salts can be made from aluminum, calcium, lithium,
magnesium, potassium, sodium and zinc. Preferred organic salts can
be made from tertiary amines and quaternary ammonium salts,
including in part, tromethamine, diethylamine,
N,N'-dibenzylethylenediamine, chloroprocaine, choline,
diethanolamine, ethylenediamine, meglumine (N-methylglucamine) and
procaine. All of these salts may be prepared by conventional means
form the corresponding compound of Formula I by reacting, for
example, the appropriate acid or base with the compound of Formula
I.
General Synthetic Procedures
[0087] The compounds of the invention can be synthesized according
to the following procedures of Schemes I-VI wherein the
R.sup.1-R.sup.3 substituents and Ar.sup.1, HetAr.sup.2 are as
defined for Formula I, above, except where further noted. 5
[0088] Scheme I shows the three step preparation of the pyrazole 5
of the present invention. In step 1, the reaction of arylmethyl
derived ketone 1 with pyridine derived aldehyde 2 either in a
solvent such as benzene or toluene in the presence of a base such
as pyridine or in a mixture of acids such as acetic acid and
hydrogen bromide gives the .alpha.,.beta.-unsaturated ketone 3. In
step 2, in the presence of base such as sodium hydroxide,
.alpha.,.beta.-unsaturated ketone 3 is converted to the
corresponding epoxide 4 by the treatment with hydrogen peroxide
solution at room temperature. In step 3, epoxide 4 is condensed
with hydrazine in a suitable solvent such as ethanol at temperature
ranging up to the boiling point to form pyrazole 5. Alternatively,
pyrazole 5 can be prepared by treatment of 3 with tosyl hydrazide
in the presence of an acid such as acetic acid at reflux. 6
[0089] Scheme II shows the synthesis of pyrazole 12 containing a
heteroaromatic ring by three routes. In Route 1, ketone 6 is
condensed with hydrazine 7 to give substituted hydrazine 9, which
is then reacted with acyl halide or anhydride 10 at low temperature
to provide acyl hydrazone 11. Upon heating at temperature up to
200.degree. C., hydrazone 11 is converted to pyrazole 12. In Route
2, acyl hydrazone 11 is formed directly by reaction of ketone 6
with acyl hydrazide 8 at room temperature. Acyl hydrazide 8 may be
formed by reaction of hydrazine with a carboxylic acid ester.
Heating 11 as above then provides pyrazole 12. In Route 3, ketone 6
is treated with acyl hydrazide 8 at from room temperature to
-200.degree. C. to give pyrazole 12 directly. Alternatively, this
condensation may be carried out in an acidic solvent, such as
acetic acid, or in a solvent containing acetic acid. 7
[0090] Cyanokerone 13 may be synthesized according to the procedure
described by I. Lantos et al in J. Org. Chem., volume 53, pp.
4223-4227 (1988) for the synthesis of the p-fluoro compound (X
p-F). This procedure, which is incorporated herein by reference,
can be used to synthesize cyanoketones such as 13 wherein X is
selected from, for example, halogen, alkyl and alkoxy. Cyanoketones
such as 13 may be converted to pyrazoles 14 by reaction with a
hydrazine in a suitable solvent, such as benzene or toluene. A
catalyst such as acetic acid may be employed. When hydrazine itself
is employed, the ring nitrogen atoms of the pyrazole thus obtained
bear no substituent except hydrogen on one of the ring nitrogen
atoms. When a substituted hydrazine, such as methylhydrazine is
employed, the product pyrazole 14 bears a substituent on the ring
nitrogen atom adjacent to the aminated ring carbon atom, as shown
in Scheme 1. The resultant aminopyrazole 14 may be acylated or
sulfonylated to form substituted aminopyrazole 15 by treatment with
a suitably activated carboxylic or sulfonic acid in a suitable
solvent such as pyridine. Examples of a suitably activated
carboxylic acid include acetic anhydride or benzoyl chloride.
Examples of a suitably activated sulfonic acid include
methanesulfonyl chloride, p-toluenesulfonyl chloride or sulfamyl
chloride. 8
[0091] Scheme IV illustrates the synthesis of
3-pyridyl-4-aryl-pyrazoles of the present invention. Benzoate 16 is
first reacted with pyridine 17 in the presence of a base, such as
an alkali metal alkoxide (preferably sodium methoxide), in a
suitable solvent, such as tetrahydrofuran. Subsequent treatment
with an acid, preferably a mineral acid such as hydrochloric acid,
yields the desoxybenzoin 18. Desoxybenzoin 18 is then converted to
ketone 19 by treatment with an excess of dimethylformamide dimethyl
acetal. Ketone 19 is then reacted with hydrazine in a suitable
solvent such as ethanol to yield a mixture of pyrazoles 20 and 21.
In Scheme IV, R.sup.4 represents one or more radicals independently
selected from the optional substituents previously defined for
Ar.sup.1; and R.sup.5 represents one or more radicals independently
selected from the optional substituents previously defined for
HetAr.sup.2.
[0092] The 3-pyrimidinyl-4-aryl-pyrazoles of the present invention
can be synthesized in the manner of Scheme IV by replacing pyridine
17 with the corresponding pyrimidine. 9
[0093] In Scheme V, hydroxyalkyl pyrazoles 22 and 23 are converted
to sulfonate derivatives by reaction with an alkyl- or arylsulfonyl
halide. These sulfonates are then reacted with ammonia or primary
amines or secondary amines to give the corresponding
1-amino-pyrazoles 24 and 25, respectively. In Scheme V, n is 1, 2,
3, 4 or 5; R.sup.4 and R.sup.5 are as defined in Scheme IV; R.sup.6
and R.sup.7 are independently selected, for example, from hydrogen,
alkyl and aryl, or together with the nitrogen atom to which they
are attached form a 4-8 membered ring that may contain one or more
additional heteroatoms selected from oxygen, nitrogen or sulfur.
10
[0094] Scheme VI is similar to Scheme IV except that desoxybenzoin
18 is first reacted with hydrazine in a suitable solvent such as
ethanol to yield hydrazine 26. Hydrazine 26 is then converted to
pyrazole 20 (rather than a mixture of pyrazoles 20 and 21 as in
Scheme IV) by treatment with an excess of dimethylformamide
dimethyl acetal. In Scheme VI, R.sup.4 and R.sup.5 are as defined
in Scheme V.
[0095] The following examples contain detailed descriptions of the
methods of preparation of compounds of Formula I. These detailed
descriptions fall within the scope, and serve to exemplify, the
above described General Synthetic Procedures which form part of the
invention. These detailed descriptions are presented for
illustrative purposes only and are not intended as a restriction on
the scope of the invention. All parts are by weight and
temperatures are in Degrees centigrade unless otherwise indicated.
All compounds showed NMR spectra consistent with their assigned
structures. In some cases, the assigned structures were confirmed
by nuclear Overhauser effect (NOE) experiments.
[0096] The following abbreviations are used:
[0097] HCl--hydrochloric acid
[0098] MgSO.sub.4--magnesium sulfate
[0099] Na.sub.2SO.sub.4--sodium sulfate
[0100] NaIO.sub.4--sodium periodate
[0101] NaHSO.sub.3--sodium bisulfite
[0102] NaOH--sodium hydroxide
[0103] KOH--potassium hydroxide
[0104] P.sub.2O.sub.5--phosphorus pentoxide
[0105] MeOH--methanol
[0106] EtOH--ethanol
[0107] HOAc (or AcOH)--acetic acid
[0108] EtOAc--ethyl acetate
[0109] H.sub.2O--water
[0110] H.sub.2O.sub.22--hydrogen peroxide
[0111] CH.sub.2Cl.sub.2--methylene chloride
[0112] NaOMe--sodium methoxide
[0113] h--hour
[0114] hr--hour
[0115] min--minutes
[0116] THF--tetrahydrofuran
[0117] TLC--thin layer chromatography
[0118] DSC--differential scanning calorimetry
[0119] b.p.--boiling point
[0120] m.p.--melting point
[0121] eq--equivalent
EXAMPLE 1
[0122] 11
[0123] 4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridine
[0124] Step 1: Preparation of
3-phenyl-4-(4-pyridyl)-3-butene-2-one
[0125] 3-Phenyl-4-(4-pyridyl)-3-butene-2-one was prepared by the
method of Reichert and Lechner, Arzneim.-Forsch. 15, 36 (1965),
which is incorporated by reference herein.
[0126] Step 2: Preparation of
3-phenyl-4-(4-pyridyl)-3,4-epoxy-2-butanone
[0127] To a stirred solution of
3-phenyl-4-(4-pyridyl)-3-butene-2-one (step 1) (500 mg, 2.24 mmol)
in methanol (10 ml) at room temperature was added an aqueous
solution (9 ml) of sodium hydroxide (100 mg, 2.24 mmol) and
hydrogen peroxide (0.5 ml of 30% aqueous solution, 4.4 mmol). After
stirring for 2 hours, sodium chloride was added and the reaction
was extracted with ethyl acetate. The combined organic layers were
dried over magnesium sulfate, filtered, and concentrated in vacuo
to provide the crude 3-phenyl-4-(4-pyridyl)-3,4-epoxy-2-butanone
(385 mg, 65%) as an oil. This was used in the next step without
further purification.
[0128] Step 3: Preparation of
4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyridin- e
[0129] A solution of 3-phenyl-4-(4-pyridyl)-3,4-epoxy-2-butanone
(step 2) (350 mg, 1.46 mmol) and anhydrous hydrazine (0.7 ml, 20
mmol) in ethanol (3 ml) was heated at reflux for 4 hours. The
reaction was cooled, and the solvent was evaporated to dryness. The
resulting residue was purified by chromatography (silica gel, 1:1
acetone/hexane) to give the desired product as a crystalline solid,
which was recrystallized from ethyl acetate and hexane to give pure
4-(3-methyl-4-phenyl-1H-pyrazol-5-yl)pyri- dine (145 mg, 42%): m.p.
164-165.degree. C.
[0130] Anal. Calc'd for C.sub.15H.sub.13N.sub.3 (235.29): C, 76.57;
H, 5.57; N, 17.86. Found: C, 76.49; H, 5.45; N, 17.70.
[0131] The compounds of Examples 2 through 8 were synthesized in
accordance with the chemistry described above (particularly in
Scheme III) by selection of the corresponding starting
reagents:
EXAMPLE 2
[0132] 12
[0133] 4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine
[0134] The cyanoketone 1 of Scheme III wherein X is p-fluoro was
synthesized according to the procedure of I. Lantos et al., J. Org.
Chem., 53, 4223-4227 (1988), which is incorporated herein by
reference. A solution of the cyanoketone (10 g, 41 mmol), hydrazine
hydrate (2.5 ml) and acetic acid (5.2 ml) in benzene (100 ml) was
refluxed for 4 hours. The reaction was cooled and extracted with 3N
HCl. The combined acid extracts were basified to pH 10 using
concentrated ammonium hydroxide with cooling. The basic aqueous
layer was extracted with methylene chloride and the combined
organic extracts were dried over magnesium sulfate.
[0135] The drying agent was filtered and the filtrate concentrated
in vacuo to give the crude
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-a- mine which was
purified by recrystallization from ethyl acetate and hexane.
Purified 4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine had
m.p. 178-180.degree. C. (capillary).
[0136] Anal. Calc'd for C.sub.14H.sub.11N.sub.4F+0.25H.sub.2O: C,
64.99; H, 4.48; N, 21.65. Found: C, 64.99; H, 4.48; N, 21.54.
EXAMPLE 3
[0137] 13
[0138]
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]methanesulfona-
mide
[0139] A solution of
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine prepared as
set forth in Example 2 (200 mg, 0.77 mmol) and methanesulfonyl
chloride (90 mg) in pyridine (5 ml) was stirred at room temperature
overnignt. The pyridine was removed in vacuo and the residue was
treated with methylene chloride and water. The resultant
precipitate was filtered to give
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl-
]methanesulfonamide. Additional
N-[4(4-fluorophenyl)-5-(4-pyridinyl)-1H-py-
razol-3-yl]methanesulfonamide was contained in the methylene
chloride layer. The methylene chloride was stripped in vacuo and
the residue purified by chromatography on silica gel using mixtures
of ethyl acetate and methanol as eluents. The purified
N-[4(4-fluorophenyl)-5-(4-pyridinyl-
)-1H-pyrazol-3-yl]methanesulfonamide had m.p. 168-170.degree.
C.
[0140] Anal. Calc'd for
C.sub.15H.sub.13N.sub.4SO.sub.2F+0.25H.sub.2O: C, 53.48; H, 4.04;
N, 16.63. Found: C, 53.41; H, 3.78; N, 16.52.
EXAMPLE 4
[0141] 14
[0142]
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylsul-
famide
[0143] Methyl sulfamyl chloride was synthesized by refluxing a
solution of methylsulfamic acid (1.0 g) in phosphorus oxychloride
(10 mL) for 6 hours. The excess phosphorus oxychloride was removed
in vacuo and the residual oil was used for the synthesis of the
product without further treatment. A solution of
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-- amine prepared as
set forth in Example 2 (200 mg, 0.77 mmol) and approximately 1 mmol
of the above oil in pyridine (5 ml) was stirred at room temperature
for 2 hours. The reaction was stripped in vacuo and the residue
purified by chromatography on silica gel using ethyl acetate and
mixtures of ethyl acetate and methanol as eluents to give
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-1-yl]-N'-methylsulfami-
de as a crystalline solid, m.p. 194-195.degree. C.
[0144] Anal. Calc'd for
C.sub.15H.sub.14N.sub.5SO.sub.2F+1.0H.sub.2O: C, 49.31; H, 4.41; N,
19.17. Found: C, 49.13; H, 3.97; N, 19.01.
EXAMPLE 5
[0145] 15
[0146] [4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]urea
[0147] A suspension of
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-ami- ne prepared as
set forth in Example 2 (200 mg, 0.77 mmol) in a solution of
di-tert-butyl carbonate (185 mg, 0.9 mmol) and
4-dimethylaminopyridine (DMAP) (10 mg) in methylene chloride (10
ml) was stirred at room temperature for 20 minutes, during which
time, the suspended material dissolved. N-Propylamine (50 mg) was
added and stirring was continued at room temperature for 1 hour.
The reaction was then refluxed for 15 minutes, cooled and stripped
in vacuo. Treatment with ethyl acetate and hexane resulted in the
deposition of crystals of the tert-butoxycarbonyl derivative, m.p.
183-184.degree. C.
[0148] Anal. Calc'd for CO.sub.9H.sub.19N.sub.4O.sub.2F: C, 64.40;
H, 5.40; N, 15.81. Found: C, 64.66; H, 5.63; N, 15.63.
[0149] A solution of the tert-butoxycarbonyl derivative above (100
mg, 0.3 mmol) in tetrahydrofuran was treated with ammonia at
80.degree. C. in a pressure bottle for 12 hours. The reaction was
stripped in vacuo and the residue was purified by chromatography on
silica gel eluting with mixtures of ethyl acetate and methanol. The
purified (4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]urea
thus obtained had m.p. 224-225.degree. C.
[0150] Anal. Calc'd for C.sub.15H.sub.12NO: C, 60.60; H, 4.07; N,
23.56. Found: C, 60.21; H, 4.11; N, 23.30.
EXAMPLE 6
[0151] 16
[0152]
[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]sulfamide
[0153] Sulfamyl chloride was synthesized from chlorosulfonyl
isocyanate according to the procedure described by R. Graf in
Chemische Berichte, p. 509 (1959), which is incorporated herein by
reference. A solution of
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine prepared as
set forth in Example 2 (200 mg, 0.77 mmol), sulfamyl chloride (100
mg, 0.8 mmol) and triethylamine (200 mg, 2 mmol) in benzene (5 ml)
and acetonitrile (5 ml) was stirred at room temperature for 2
hours. The reaction was stripped in vacuo and residue was treated
with water and basified to pH 7 with ammonium hydroxide. The
resultant precipitate was purified by chromatography on silica gel
using mixtures of ethyl acetate and methanol as eluents. The
purified [4-(4-fluorophenyl)-5-(4-pyridinyl)-
-1H-pyrazol-3-yl]sulfamide thus obtained had m.p. 201-202.degree.
C.
[0154] Anal. Calc'd for C.sub.14H.sub.12N.sub.5SO.sub.2F: C, 50.44;
H, 3.63; N, 21.01. Found: C, 50.43; H, 3.45; N, 20.89.
EXAMPLE 7
[0155] 17
[0156]
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine
[0157] A solution of cyanoketone 1 of Scheme III wherein X is
p-chloro (1.5 g, 5.19 mmol), methylhydrazine (0.35 ml) and acetic
acid (0.75 ml) in benzene (15 ml) was refluxed for 3.5 hours. The
reaction was cooled and extracted with 3N HCl. The aqueous layer
was concentrated on the rotary evaporator and then basified with
ammonium hydroxide. The resultant precipitate was recrystallized
from methanol to give pure
4-(4-chlorophenyl)-1-methyl-3-(4-pyridinyl)-1H-pyrazol-5-amine,
m.p. 257-258.degree. C.
[0158] Anal. Calc'd for C.sub.15H.sub.13N.sub.4Cl: C, 63.27; H,
4.60; N, 19.68. Found: C, 62.93; H, 4.45; N, 19.64.
EXAMPLE 8
[0159] 18
[0160]
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-yl]-N'-methylure-
a
[0161] A solution of
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-3-amine prepared as
set forth in Example 2 (100 mg, 0.38 mmol), methyl isocyanate (22
mg, 0.39 mmol) and 4-dimethylaminopyridine (2.5 mg) in methylene
chloride (10 ml) was stirred at room temperature for 30 minutes.
The reaction was stripped in vacuo. The residue was triturated with
hexane and the solid filtered to give pure
N-[4-(4-fluorophenyl)-5-(4-pyridinyl)-
-1H-pyrazol-3-yl]-N'-methylurea, m.p. 212-213.degree. C.
[0162] Anal. Calc'd for C.sub.16H.sub.14N.sub.5FO: C, 61.73; H,
4.53; N, 22.50. Found: C, 61.63; H, 4.55; N, 22.47.
[0163] The compounds of Examples 9 through 11 were synthesized in
accordance with the chemistry described above (particularly in
Scheme IV) by selection of the corresponding starting reagents:
EXAMPLE 9
[0164] 19
[0165] 4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine
[0166] Step 1
[0167] Methyl isonicotinate (13.7 g, 0.1 mole) and ethyl
4-fluorophenylacetate (18.2 g, 0.1 mole) were mixed together, then
sodium methoxide (8.1 g, 0.15 mole) was added. The mixture was
heated to 60-70.degree. C. for 24 hours while nitrogen was blown
through the flask to eliminate methanol. Concentrated hydrochloric
acid (50 mL) then was added and the reaction mixture was refluxed
for 3 hours. After addition of water (30 mL), the reaction mixture
was extracted with chloroform, and the water phase was neutralized
to pH 6-7 with aqueous sodium hydroxide (1M). The precipitate
formed was collected by filtration, washed with water and dried
under vacuum to give 10 g of 2-(4-fluorophenyl)-1-(4'-pyr-
idyl)-ethan-1-one (yield: 46%). .sup.1H NMR: consistent with the
assigned structure and/or its tautomer.
[0168] Step 2
[0169] 2-(4-fluorophenyl)-1-(4'-pyridyl)-ethan-1-one prepared above
(1 g) was dissolved in 50 mL tetrahydrofuran and
N,N-dimethylformamide dimethyl acetal (5 mL) was added. The mixture
was stirred at room temperature for 2 days. After evaporating the
solvent, the solid obtained was washed with hexane and 1 g of the
vinyl amine was obtained. This vinyl amine (0.5 g) was dissolved in
ethanol (15 mL) and hydrazine hydrate (5 mL) was added. The mixture
was stirred at 0.degree. C. for 2 hours and then evaporated to
dryness. After recrystallization from methanol/water, 400 mg of
4-[4-(4-fluorophenyl)-1H-pyrazol-3-yl]pyridine was obtained in 91%
yield. MS, 240(M+1); .sup.1H NMR: consistent with the assigned
structure; Anal. Calc'd for C.sub.14H.sub.10FN.sub.3.0.2H.sub.2O:
C, 69.24; H, 4.32; N, 17.30. Found: C, 69.54; H, 4.06; N,
17.43.
EXAMPLE 10
[0170] 20
[0171] 4-[4-(4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine
[0172] When methylhydrazine was substituted for hydrazine hydrate
in Step 2 of Example 9,
4-[4-(4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine (the
N-methyl derivative corresponding to the compound of Example 9) was
obtained. Purification by recrystallization from toluene and hexane
give the pure
4-[4-(4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine in 57%
yield. MS m/z: 254 (M+1). .sup.1H NMR: consistent with the assigned
structure. Anal. calc'd for C.sub.15H.sub.12FN.sub.3: C, 71.13; H,
4.78; N, 16.69. Found: C, 70.99; H, 4.68; N, 16.65.
EXAMPLE 11
[0173] 21
[0174] 4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazole-1-ethanol and
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazole-1-ethanol
[0175] The procedure set forth in Example 9 was followed except
that 2-hydroxyethyl hydrazine was substituted for hydrazine
hydrate. 4-(4-Fluorophenyl)-3-(4-pyridinyl)-1H-pyrazole-1-ethanol
and 4-(4-fluorophenyl)-5(4-pyridinyl)-1H-pyrazole-1-ethanol were
obtained as a mixture by recrystallization from toluene and hexane
in 67% yield. .sup.1H NMR: consistent with the assigned
structure.
[0176] Mass spectrum, m/z: 284 (M+1). Anal. calc'd for
C.sub.16H.sub.14FN.sub.3O: C, 67.83; H, 4.98; N, 14.83. Found: C,
67.86; H, 5.04; N, 14.85.
[0177] The compounds of Examples 12 and 13 were synthesized in
accordance with the chemistry described above (particularly in
Scheme V) by selection of the corresponding starting reagents:
EXAMPLE 12
[0178] 22
[0179]
4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazole-1-ethan-
amine and
4-(4-fluorophenyl)-N,N-dimethyl-s-(4-pyridinyl)-1H-pyrazole-1-et-
hanamine
[0180] 4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazole-1-ethanol (or
4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazole-1-ethanol) prepared
as set forth in Example 11 (1.36 g) was dissolved in 30 mL pyridine
and cooled to 0.degree. C., whereupon methanesulfonyl chloride (0.6
mL) was added. After stirring at 0.degree. C. for 12 hours, about
20 g of ice was added, and the mixture was extracted with toluene
(300 ml). After evaporation, the residue was used directly without
further purification. 0.7 g of the above obtained compound was
dissolved in methanol (25 mL), and dimethylamine/THF solution (4M,
2 mL) was added. The reaction mixture was refluxed for 12 hours,
then evaporated to dryness. The residue was purified by
chromatography (methanol/dichloromethane 1:10). A mixture (0.59 g)
of 4-(4-fluorophenyl)-N,N-dimethyl-3-(4-pyridinyl)-1H-pyrazole-1-
-ethanamine and
4-(4-fluorophenyl)-N,N-dimethyl-5-(4-pyridinyl)-1H-pyrazol-
e-1-ethanamine were obtained. .sup.1H NMR: consistent with the
assigned structure. Mass spectrum, m/z: 311 (M+1). Anal. calc'd
C.sub.18H.sub.19N.sub.4F.0.55H.sub.2O: C, 67.50; H, 6.33; N, 17.49.
Found: C, 67.21; H, 6.46; N, 17.14.
EXAMPLE 13
[0181] 23
[0182]
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]ethyl]
morpholine and
4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-1-yl]e-
thyl]morpholine
[0183] The procedure set forth in Example 11 was followed, except
that morpholine was substituted for dimethylamine, to produce a
mixture of
4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]ethyl]morpholine
and
4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazol-1-yl]ethyl]morpho-
line. Mass spectrum, m/z: 353 (M+1). Anal. calc'd for
C.sub.20H.sub.21N.sub.4OF+0.5H.sub.2O: C, 66.47; H, 6.14; N, 15.50.
Found: C, 66.57; H, 6.27; N, 15.14.
[0184] The compound of Example 14 was synthesized in accordance
with the chemistry described above (particularly in Scheme VI) by
selection of the corresponding starting reagents:
EXAMPLE 14
[0185] 24
[0186] 4-[4-4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine
[0187] 2-(4-fluorophenyl)-1-(4'-pyridyl)-ethan-1-one prepared as
set forth in step 1 of Example 9 (0.5 g, 0.00232 moles) was mixed
with of 98% methyl hydrazine (0.2 g, 0.00462 moles) in 10 mL of
ethanol containing 0.1 mL of acetic acid in a 50 mL Erlenmeyer
flask. After gentle boiling (30 minutes on a steam bath) a small
sample was evacuated at high vacuum and examined by NMR to confirm
completion of hydrazone formation. The reaction mixture was
concentrated to a pasty mass and 3.6 mL of DMF dimethylacetal
(0.027 moles) was then added and heated to 80.degree. C. for 30
minutes, at which point a clear yellow viscous solution was
obtained. The reaction was checked for completion (TLC or NMR) and
concentrated and taken up in 20 mL of chloroform. After washing
with water (10 mL), the organic layer was extracted with 15 mL of
10% HCl. The water layer was then treated with 0.5 g of activated
charcoal at 70.degree. C. for 10 minutes, filtered through celite,
neutralized cautiously to pH 7-8 with vigorous stirring and
cooling. The fine off-white precipitate was filtered and dried. NMR
was found to be in agreement with the proposed structure. The
precipitate,
4-[4-(4-fluorophenyl)-1-methyl-1H-pyrazol-3-yl]pyridine, obtained
in quantitative yield was filtered, washed with ether and dried.
Yield: 0.45 g (77%). Mass spectrum, m/z: 254. Anal. calc'd: C,
62.18; H, 4.52; N, 14.50. Found: C, 62.39; H, 4.07; N, 14.24.
EXAMPLE 15
[0188] 25
[0189] 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine
[0190] 4-[4-(4-chlorophenyl)-1H-pyrazol-3-yl]pyridine was prepared
according to the procedure set forth in Example 9 except that ethyl
4-chlorophenylacetate was substituted for ethyl
4-fluorophenylacetate; m.p. 204-207.degree. C.
[0191] Anal. Calc'd: C, 65.76; H, 3.94; N, 16.43. Found: C, 65.44;
H, 3.78; N, 16.04.
EXAMPLE 16
[0192] 26
[0193] 4-(4-phenyl-1H-pyrazol-5-yl)pyridine
[0194] 4-(4-phenyl-1H-pyrazol-5-yl)pyridine can be prepared in
accordance with the procedure set forth in Example 9 by
substituting ethylphenylacetate for ethyl
4-fluorophenylacetate.
EXAMPLE 17
[0195] 27
[0196]
1-methyl-4-[2-[4-(4-fluorophenyl)-3-(4-pyridinyl)-1H-pyrazol-1-yl]]-
piperidine and
1-methyl-4-[2-[4-(4-fluorophenyl)-5-(4-pyridinyl)-1H-pyrazo-
l-1-yl]piperidine
[0197] This compound can be prepared using the procedure set forth
for the synthesis of the compound of Example 11 by substituting
4-hydrazino-N-methylpiperidine for hydroxyethyl hydrazine.
4-Hydrazino-N-methylpiperidine is synthesized as disclosed in
Ebnoether et al, Helv. Chim. Acta (1959) 42, 533, 541, 560. The
resulting mixture is separated into the respective pure title
compounds by chromatography on silica gel, eluting with
methanol/dichloromethane (1:10), or other suitable solvent
system.
BIOLOGICAL EVALUATION
[0198] p38 Kinase Assay
[0199] Cloning of Human p38a:
[0200] The coding region of the human p38a cDNA was obtained by
PCR-amplification from RNA isolated from the human monocyte cell
line THP.1. First strand cDNA was synthesized from total RNA as
follows: 2 .mu.g of RNA was annealed to 100 ng of random hexamer
primers in a 10 .mu.l reaction by heating to 70.degree. C. for 10
minutes followed by 2 minutes on ice. cDNA was then synthesized by
adding 1 .mu.l of RNAsin (Promega, Madison Wis.), 2 .mu.l of 50 mM
dNTP's, 4 .mu.l of 5.times. buffer, 2 .mu.l of 100 mM DTT and 1
.mu.l (200 U) of Superscript II.TM. AMV reverse transcriptase.
Random primer, dNTP's and Superscript.TM. reagents were all
purchased from Life-Technologies, Gaithersburg, Mass. The reaction
was incubated at 42.degree. C. for 1 hour. Amplification of p38
cDNA was performed by aliquoting 5 .mu.l of the reverse
transcriptase reaction into a 100 .mu.l PCR reaction containing the
following: 80 .mu.l dH.sub.2O, 2 .mu.l 50 mM dNTP's, 1 .mu.l each
of forward and reverse primers (50 pmol/.mu.l), 10 .mu.l of
10.times. buffer and 1 .mu.l Expand.TM. polymerase (Boehringer
Mannheim). The PCR primers incorporated Bam HI sites onto the 5'
and 3' end of the amplified fragment, and were purchased from
Genosys. The sequences of the forward and reverse primers were
5'-GATCGAGGATTCATGTCTCAGGAGAGGCCCA-3' and
5'GATCGAGGATTCTCAGGACTCCAT- CTCTTC-3' respectively. The PCR
amplification was carried out in a DNA Thermal Cycler (Perkin
Elmer) by repeating 30 cycles of 94.degree. C. for 1 minute,
60.degree. C. for 1 minute and 68.degree. C. for 2 minutes. After
amplification, excess primers and unincorporated dNTP's were
removed from the amplified fragment with a Wizard.TM. PCR prep
(Promega) and digested with Bam HI (New England Biolabs). The Bam
HI digested fragment was ligated into BamHI digested pGEX 2T
plasmid DNA (PharmaciaBiotech) using T-4 DNA ligase (New England
Biolabs) as described by T. Maniatis, Molecular Cloning: A
Laboratory Manual, 2nd ed. (1989). The ligation reaction was
transformed into chemically competent E. coli DH10B cells purchased
from Life-Technologies following the manufacturer's instructions.
Plasmid DNA was isolated from the resulting bacterial colonies
using a Promega Wizard.TM. miniprep kit. Plasmids containing the
appropriate Bam HI fragment were sequenced in a DNA Thermal Cycler
(Perkin Elmer) with Prism.TM. (Applied Biosystems Inc.). cDNA
clones were identified that coded for both human p38a isoforms (Lee
et al. Nature 372, 739). One of the clones which contained the cDNA
for p38a-2 (CSBP-2) inserted in the cloning site of pGEX 2T, 3' of
the GST coding region was designated pMON 35802. The sequence
obtained for this clone is an exact match of the cDNA clone
reported by Lee et al. This expression plasmid allows For the
production of a GST-p38a fusion protein.
[0201] Expression of Human p38a:
[0202] GST/p38a fusion protein was expressed from the plasmid pMON
35802 in E. coli, stain DH10B (Life Technologies, Gibco-BRL).
Overnight cultures were grown in Luria Broth (LB) containing 100
mg/ml ampicillin. The next day, 500 ml of fresh LB was inoculated
with 10 ml of overnight culture, and grown in a 2 liter flask at
37.degree. C. with constant shaking until the culture reached an
absorbance of 0.8 at 600 nm. Expression of the fusion protein was
induced by addition of isopropyl b-D-thiogalactosidse (IPTG) to a
final concentration of 0.05 mM. The cultures were shaken for three
hours at room temperature, and the cells were harvested by
centrifugation. The cell pellets were stored frozen until protein
purification.
[0203] Purification of p38 Kinase-.alpha.:
[0204] All chemicals were from Sigma Chemical Co. unless noted.
Twenty grams of E. coli cell pellet collected from five 1 L shake
flask fermentations was resuspended in a volume of PBS (140 mM
NaCl, 2.7 mM KCl, 10 mM Na.sub.2HPO.sub.4, 1.8 mM KH.sub.2PO.sub.4,
pH 7.3) up to 200 ml. The cell suspension was adjusted to 5 mM DTT
with 2 M DTT and then split equally into five 50 ml Falcon conical
tubes. The cells were sonnicated (Ultrasonics model W375) with a 1
cm probe for 3.times.1 minutes (pulsed) on ice. Lysed cell material
was removed by centrifugation (12,000.times.g, 15 minutes) and the
clarified supernatant applied to glutathione-sepharose resin
(Pharmacia).
[0205] Glutathione-Sepharose Affinity Chromatography:
[0206] Twelve ml of a 50% glutathione sepharose-PBS suspension was
added to 200 ml clarified supernatant and incubated batchwise for
30 minutes at room temperature. The resin was collected by
centrifugation (600.times.g, 5 min) and washed with 2.times.150 ml
PBS/1% Triton X-100, followed by 4.times.40 ml PBS. To cleave the
p38 kinase from the GST-p38 fusion protein, the
glutathione-sepharose resin was resuspended in 6 ml PBS containing
250 units thrombin protease (Pharmacia, specific activity >7500
units/mg) and mixed gently for 4 hours at room temperature. The
glutathione-sepharose resin was removed by centrifugation
(600.times.g, 5 min) and washed 2.times.6 ml with PBS. The PBS wash
fractions and digest supernatant containing p38 kinase protein were
pooled and adjusted to 0.3 mM PMSF.
[0207] Mono Q Anion Exchange Chromatography:
[0208] The thrombin-cleaved p38 kinase was further purified by
FPLC-anion exchange chromatography. Thrombin-cleaved sample was
diluted 2-fold with Buffer A (25 mM HEPES, pH 7.5, 25 mM
beta-glycerophosphate, 2 mM DTT, 5% glycerol) and injected onto a
Mono Q HR 10/10 (Pharmacia) anion exchange column equilibrated with
Buffer A. The column was eluted with a 160 ml 0.1 M-0.6 M
NaCl/Buffer A gradient (2 ml/minute flowrate). The p38 kinase peak
eluting at 200 mM NaCl was collected and concentrated to 3-4 ml
with a Filtron 10 concentrator (Filtron Corp.).
[0209] Sechacryl S100 Gel Filtration Chromatography:
[0210] The concentrated Mono Q-p38 kinase purified sample was
purified by gel filtration chromatography (Pharmacia HiPrep 26/60
Sephacryl S100 column equilibrated with Buffer B (50 mM HEPES, pH
7.5, 50 mM NaCl, 2 mM DTT, 5% glycerol)). Protein was eluted from
the column with Buffer B at a 0.5 ml/minute flowrate and protein
was detected by absorbance at 280 nm. Fractions containing p38
kinase (detected by SDS-polyacrylamide gel electropnhoresis) were
pooled and frozen at -80.degree. C. Typical purified protein yields
from 5 L E. coli shake flasks fermentations were 35 mg p38
kinase.
[0211] In Vitro Assay
[0212] The ability of compounds to inhibit human p38 kinase alpha
was evaluated using two in vitro assay methods. In the first
method, activated human p38 kinase alpha phosphorylates a
biotinylated substrate, PHAS-I (phosphorylated heat and acid stable
protein-insulin inducible), in the presence of gamma .sup.32P-ATP
(.sup.32P-ATP). PHAS-I was biotinylated prior to the assay and
provides a means of capturing the substrate which is phosphorylated
during the assay. p38 Kinase was activated by MKK6. Compounds were
tested in 10 fold serial dilutions over the range of 100 .mu.M to
0.001 .mu.M using 1% DMSO. Each concentration of inhibitor was
tested in triplicate.
[0213] All reactions were carried out in 96 well polypropylene
plates. Each reaction well contained 25 mM HEPES pH 7.5, 10 mM
magnesium acetate and 50 .mu.M unlabeled ATP. Activation of p38 was
required to achieve sufficient signal in the assay. Biotinylated
PHAS-I was used at 1-2 .mu.g per 50 .mu.l reaction volume, with a
final concentration of 1.5 .mu.M. Activated human p38 kinase alpha
was used at 1 .mu.g per 50 .mu.l reaction volume representing a
final concentration of 0.3 .mu.M. Gamma .sup.32P-ATP was used to
follow the phosphorylation of PHAS-I. .sup.32P-ATP has a specific
activity of 3000 Ci/mmol and was used at 1.2 .mu.Ci per 50 .mu.l
reaction volume. The reaction proceeded either for one hour or
overnight at 30.degree. C.
[0214] Following incubation, 20 .mu.l of reaction mixture was
transferred to a high capacity streptavidin coated filter place
(SAM-streptavidin-matrix, Promega) prewetted with phosphate
buffered saline. The transferred reaction mix was allowed to
contact the streptavidin membrane of the Promega plate for 1-2
minutes. Following capture of bictinylated PHAS-I with .sup.32P
incorporated, each well was washed to remove unincorporated
.sup.32P-ATP three times with 2M NaCl, three washes of 2M NaCl with
1% phosphoric, three washes of distilled water and finally a single
wash of 95% ethanol. Filter plates were air dried and 20 .mu.l of
scintillant was added. The plates were sealed and counted.
[0215] A second assay format was also employed that is based on p38
kinase alpha induced phosphorylation of EGFRP (epidermal growth
factor receptor peptide, a 21 mer) in the presence of .sup.33P-ATP.
Compounds were tested in 10 fold serial dilutions over the range of
100 .mu.M to 0.001 .mu.M in 1% DMSO. Each concentration of
inhibitor was tested in triplicate. Compounds were evaluated in 501
reaction volumes in the presence of 25 mM Hepes pH 7.5, 10 mM
magnesium acetate, 4% glycerol, 0.4% bovine serum albumin, 0.4 mM
DTT, 50 .mu.M unlabeled ATP, 25 .mu.g EGFRP (200 .mu.M), and 0.05
uCi gamma .sup.33P-ATP. Reactions were initiated by addition of
0.09 .mu.g of activated, purified human GST-p38 kinase alpha.
Activation was carried out using GST-MKK6 (5:1, p38:MKK6) for one
hour at 30.degree. C. in the presence of 50 .mu.M ATP. Following
incubation for 60 minutes at room temperature, the reaction was
stopped by addition of 1501 of AG 1.times.8 resin in 900 mM sodium
formate buffer, pH 3.0 (1 volume resin to 2 volumes buffer). The
mixture was mixed three times with pipetting and the resin was
allowed to settle. A total of 50 .mu.l of clarified solution head
volume was transferred from the reaction wells to Microlite-2
plates. 150 .mu.l of Microscint 40 was then added to each well of
the Microlite plate, and the plate was sealed, mixed, and
counted.
[0216] TNF Cell Assays
[0217] Method of Isolation of Human Peripheral Blood Mononuclear
Cells:
[0218] Human whole blood was collected in Vacutainer tubes
containing EDTA as an anticoagulant. A blood sample (7 ml) was
carefully layered over 5 ml PMN Cell Isolation Medium (Robbins
Scientific) in a 15 ml round bottom centrifuge tube. The sample was
centrifuged at 450-500.times.g for 30-35 minutes in a swing out
rotor at room temperature. After centrifugation, the top band of
cells were removed and washed 3 times with PBS w/o calcium or
magnesium. The cells were centrifuged at 400.times.g for 10 minutes
at room temperature. The cells were resuspended in Macrophage Serum
Free Medium (Gibco BRL) at a concentration of 2 million
cells/ml.
[0219] LPS Stimulation of Human PBMs:
[0220] PBM cells (0.1 ml, 2 million/ml) were co-incubated with 0.1
ml compound (10-0.41 .mu.M, final concentration) for 1 hour in flat
bottom 96 well microtiter plates. Compounds were dissolved in DMSO
initially and diluted in TCM for a final concentration of 0.1%
DMSO. LPS (Calbiochem, 20 ng/ml, final concentration) was then
added at a volume of 0.010 ml. Cultures were incubated overnight at
37.degree. C. Supernatants were then removed and tested by ELISA
for TNF-a and IL1-b. Viability was analyzed using MTS. After 0.1 ml
supernatant was collected, 0.020 ml MTS was added to remaining 0.1
ml cells. The cells were incubated at 37.degree. C. for 2-4 hours,
then the O.D. was measured at 490-650 nM.
[0221] Maintenance and Differentiation of the U937 Human
Histiocytic Lymphoma Cell Line:
[0222] U937 cells (ATCC) were propagated in RPMI 1640 containing
10% fecal bovine serum, 100 IU/ml penicillin, 100 .mu.g/ml
streptomycin, and 2 mM glutamine (Gibco). Fifty million cells in
100 ml media were induced to terminal monocytic differentiation by
24 hour incubation with 20 ng/ml phorbol 12-myristate 13-acetate
(Sigma). The cells were washed by centrifugation (200.times.g for 5
min) and resuspended in 100 ml fresh medium. After 24-48 hours, the
cells were harvested, centrifuged, and resuspended in culture
medium at 2 million cells/ml.
[0223] LPS Stimulation of TNF Production by U937 Cells:
[0224] U937 cells (0.1 ml, 2 million/ml) were incubated with 0.1 ml
compound (0.004-50 .mu.M, final concentration) for 1 hour in 96
well microtiter plates. Compounds were prepared as 10 mM stock
solutions in DMSO and diluted in culture medium to yield a final
DMSO concentration of 0.1% in the cell assay. LPS (E coli, 100
ng/ml final concentration) was then added at a volume of 0.02 ml.
After 4 hour incubation at 37.degree. C., the amount of TNF-.alpha.
released in the culture medium was quantitated by ELISA. Inhibitory
potency is expressed as ICSO (.mu.M).
[0225] Rat Assay
[0226] The efficacy of the novel compounds in blocking the
production of TNF also was evaluated using a model based on rats
challenged with LPS. Male Harlen Lewis rats [Sprague Dawley Co.]
were used in this model. Each rat weighed approximately 300 g and
was fasted overnight prior to testing. Compound administration was
typically by oral gavage (although intraperitoneal, subcutaneous
and intravenous administration were also used in a few instances) 1
to 24 hours prior to the LPS challenge. Rats were administered 30
.mu.g/kg LPS [salmonella typhosa, Sigma Co.] intravenously via the
tail vein. Blood was collected via heart puncture 1 hour after the
LPS challenge. Serum samples were stored at -20.degree. C. until
quantitative analysis of TNF-.alpha. by Enzyme
Linked-Immuno-Sorbert Assay ("ELISA") [Biosource]. Additional
details of the assay are set forth in Perretti, M., et al., Br. J.
Pharmacol. (1993), 110, 868-874, which is incorporated by reference
in this application.
[0227] Mouse Assay
[0228] Mouse Model Of LPS-Induced TNF Aloha Production:
[0229] TNF alpha was induced in 10-12 week old BALB/c female mice
by tail vein injection with 100 ng lipopolysaccharide (from S.
Typhosa) in 0.2 ml saline. One hour later mice were bled from the
retroorbital sinus and TNF concentrations in serum from clotted
blood were quantified by ELISA. Typically, peak levels of serum TNF
ranged from 2-6 ng/ml one hour after LPS injection.
[0230] The compounds tested were administered to fasted mice by
oral gavage as a suspension in 0.2 ml of 0.5% methylcellulose and
0.025% Tween 20 in water at 1 hour or 6 hours prior to LPS
injection. The 1 hour protocol allowed evaluation of compound
potency at Cmax plasma levels whereas the 6 hour protocol allowed
estimation of compound duration of action. Efficacy was determined
at each time point as percent inhibition of serum TNF levels
relative to LPS injected mice that received vehicle only.
[0231] Induction and Assessment of Collagen-Induced Arthritis in
Mice:
[0232] Arthritis was induced in mice according to the procedure set
forth in J. M. Stuart, Collagen Autoimmune Arthritis, Annual Rev.
Immunol. 2:199 (1984), which is incorporated herein by reference.
Specifically, arthritis was induced in 8-12 week old DBA/1 male
mice by injection of 50 .mu.g of chick type II collagen (CII)
(provided by Dr. Marie Griffiths, Univ. of Utah, Salt Lake City,
Utah) in complete Freund's adjuvant (Sigma) on day 0 at the base of
the tail. Injection volume was 100 .mu.l. Animals were boosted on
day 21 with 50 .mu.g of CII in incomplete Freund's adjuvant (100
.mu.l volume). Animals were evaluated several times each week for
signs of arthritis. Any animal with paw redness or swelling was
counted as arthritic. Scoring of arthritic paws was conducted in
accordance with the procedure set forth in Wooley et al., Genetic
Control of Type II Collagen Induced Arthritis in Mice: Factors
Influencing Disease Suspectibility and Evidence for Multiple MHC
Associated Gene Control., Trans. Proc., 15:180 (1983). Scoring of
severity was carried out using a score of 1-3 for each paw (maximal
score of 12/mouse). Animals displaying any redness or swelling of
digits or the paw were scored as 1. Gross swelling of the whole paw
or deformity was scored as 2. Ankylosis of joints was scored as 3.
Animals were evaluated for 8 weeks. 8-10 animals per group were
used.
[0233] Preparation and Administration of Compounds:
[0234] The compounds tested on mice having collagen-induced
arthritis were prepared as a suspension in 0.5% methylcelluose
(Sigma, St. Louis, Mo.), 0.025% Tween 20 (Sigma). The compound
suspensions were administered by oral gavage in a volume of 0.1 ml
b.i.d. Administration began on day 20 post collagen injection and
continued daily until final evaluation on day 56. Scoring of
arthritic paws was conducted as set forth above.
[0235] Results obtained using the above-described assays are set
forth in Table I below. p38 assay and U937 cell assay results are
expressed as IC.sub.50 (.mu.m). Mouse-LPS assay results are
expressed as percent inhibition.
1TABLE I mLPS P38.alpha..sup.1 p38.alpha..sup.2 U937 (6 h @ Example
(.mu.M) (.mu.M) (.mu.M) (30 mpk) 1 30.00 13.35 10.00 2 6.21 10.61 3
2.55 >10.00 4 0.23 4.70 54 5 1.98 5.53 6 10.00 7 5.48 10.00 8
10.00 9 2.44 3.46 0.6474 42 10 7.23 0.4 1.5987 76 11 0.695 10 40 12
0.941 10 -5 13 0.86 >10 22 15 5.9 0.75 32 .sup.1p38.alpha. in
vitro results based on PHAS-I assay procedure .sup.2p38.alpha. in
vitro results based on EGFRP assay procedure
[0236] Also embraced within this invention is a class of
pharmaceutical compositions comprising the active compounds of this
invention in association with one or more non-toxic,
pharmaceutically-acceptable carriers and/or diluents and/or
adjuvants (collectively referred to herein as "carrier" materials)
and, if desired, other active ingredients. The active compounds of
the present invention may be administered by any suitable route,
preferably in the form of a pharmaceutical composition adapted to
such a route, and in a dose effective for the treatment intended.
The active compounds and composition may, for example, be
administered orally, intravascularly (IV), intraperitoneally,
subcutaneously, intramuscularly (IM) or topically.
[0237] For oral administration, the pharmaceutical composition may
be in the form of, for example, a tablet, hard or soft capsule,
lozenges, dispensable powders, suspension or liquid. The
pharmaceutical composition is preferably made in the form of a
dosage unit containing a particular amount of the active
ingredient. Examples of such dosage units are tablets or capsules.
The active ingredient may also be administered by injection (IV,
IM, subcutaneous or jet) as a composition wherein, for example,
saline, dextrose, or water may be used as a suitable carrier. The
pH of the composition may be adjusted, if necessary, with suitable
acid, base, or buffer. Suitable bulking, dispersing, wetting or
suspending agents, including mannitol and PEG 400, may also be
included in the composition. A suitable parenteral composition can
also include a compound formulated as a sterile solid substance,
including lyophilized powder, in injection vials. Aqueous solution
can be added to dissolve the compound prior to injection. The
amount of therapeutically active compounds that are administered
and the dosage regimen for treating a disease condition with the
compounds and/or compositions of this invention depends on a
variety of factors, including the age, weight, sex and medical
condition of the subject, the severity of the inflammation or
inflammation related disorder, the route and frequency of
administration, and the particular compound employed, and thus may
vary widely. The pharmaceutical compositions may contain active
ingredients in the range of about 0.1 to 1000 mg, preferably in the
range of about 7.0 to 350 mg. A daily dose of about 0.01 to 100
mg/kg body weight, preferably between about 0.1 and about 50 mg/kg
body weight and most preferably between about 0.5 to 30 mg/kg body
weight, may be appropriate. The daily dose can be administered in
one to four doses per day. In the case of skin conditions, it may
be preferable to apply a topical preparation of compounds of this
invention to the affected area two to four times a day. For
disorders of the eye or other external tissues, e.g., mouth and
skin, the formulations are preferably applied as a topical gel,
spray, ointment or cream, or as a suppository, containing the
active ingredients in a total amount of, for example, 0.075 to 30%
w/w, preferably 0.2 to 20% w/w and most preferably 0.4 to 15% w/w.
When formulated in an ointment, the active ingredients may be
employed with either paraffinic or a water-miscible ointment base.
Alternatively, the active ingredients may be formulated in a cream
with an oil-in-water cream base. If desired, the aqueous phase of
the cream base may include, for example at least 30% w/w of a
polyhydric alcohol such as propylene glycol, butane-1,3-diol,
mannitol, sorbitol, glycerol, polyethylene glycol and mixtures
thereof. The topical formulation may desirably include a compound
which enhances absorption or penetration of the active ingredient
through the skin or other affected areas. Examples of such dermal
penetration enhancers include dimethylsulfoxide and related
analogs. The compounds of this invention can also be administered
by a transdermal device. Preferably topical administration will be
accomplished using a patch either of the reservoir and porous
membrane type or of a solid matrix variety. In either case, the
active agent is delivered continuously from the reservoir or
microcapsules through a membrane into the active agent permeable
adhesive, which is in contact with the skin or mucosa of the
recipient. If the active agent is absorbed through the skin, a
controlled and predetermined flow of the active agent is
administered to the recipient. In the case of microcapsules, the
encapsulating agent may also function as the membrane. The
transdermal patch may include the compound in a suitable solvent
system with an adhesive system, such as an acrylic emulsion, and a
polyester patch. The oily phase of the emulsions of this invention
may be constituted from known ingredients in a known manner. While
the phase may comprise merely an emulsifier, it may comprise a
mixture of at least one emulsifier with a fat or an oil or with
both a fat and an oil. Preferably, a hydrophilic emulsifier is
included together with a lipophilic emulsifier which acts as a
stabilizer. It is also preferred to include both an oil and a fat.
Together, the emulsifier(s) with or without stabilizer(s) make-up
the so-called emulsifying wax, and the wax together with the oil
and fat make up the so-called emulsifying ointment base which forms
the oily dispersed phase of the cream formulations. Emulsifiers and
emulsion stabilizers suitable for use in the formulation of the
present invention include Tween 60, Span 80, cetostearyl alcohol,
myristyl alcohol, glyceryl monostearate, and sodium lauryl sulfate,
among others. The choice of suitable oils or fats for the
formulation is based on achieving the desired cosmetic properties,
since the solubility of the active compound in most oils likely to
be used in pharmaceutical emulsion formulations is very low. Thus,
the cream should preferably be a non-greasy, non-staining and
washable product with suitable consistency to avoid leakage from
tubes or other containers. Straight or branched chain, mono- or
dibasic alkyl esters such as di-isoadipate, isocetyl stearate,
propylene glycol diester of coconut fatty acids, isopropyl
myristate, decyl oleate, isopropyl palmitate, butyl stearate,
2-ethylhexyl palmitate or a blend of branched chain esters may be
used. These may be used alone or in combination depending on the
properties required. Alternatively, high melting point lipids such
as white soft paraffin and/or liquid paraffin or other mineral oils
can be used.
[0238] Formulations suitable for topical administration to the eye
also include eye drops wherein the active ingredients are dissolved
or suspended in suitable carrier, especially an aqueous solvent for
the active ingredients. The antiinflammatory active ingredients are
preferably present in such formulations in a concentration of 0.5
to 20%, advantageously 0.5 to 10% and particularly about 1.5% w/w.
For therapeutic purposes, the active compounds of this combination
invention are ordinarily combined with one or more adjuvants
appropriate to the indicated route of administration. If
administered per os, the compounds may be admixed with lactose,
sucrose, starch powder, cellulose esters of alkanoic acids,
cellulose alkyl esters, talc, stearic acid, magnesium stearate,
magnesium oxide, sodium and calcium salts of phosphoric and
sulfuric acids, gelatin, acacia gum, sodium alginate,
polyvinylpyrrolidone, and/or polyvinyl alcohol, and then tableted
or encapsulated for convenient administration. Such capsules or
tablets may contain a controlled-release formulation as may be
provided in a dispersion of active compound in hydroxypropylmethyl
cellulose. Formulations for parenteral administration may be in the
form of aqueous or non-aqueous isotonic sterile injection solutions
or suspensions. These solutions and suspensions may be prepared
from sterile powders or granules having one or more of the carriers
or diluents mentioned for use in the formulations for oral
administration. The compounds may be dissolved in water,
polyethylene glycol, propylene glycol, ethanol, corn oil,
cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium
chloride, and/or various buffers. Other adjuvants and modes of
administration are well and widely known in the pharmaceutical
art.
[0239] All patent documents listed herein are incorporated by
reference. Although this invention has been described with respect
to specific embodiments, the details of these embodiments are not
to be construed as limitations.
* * * * *